Hydrophobic interactions within biofilms of nitrifying and denitrifying bacteria in biofilters

Hydrophobicity of the solid surface and microbial cell surface is important factor for the development of biofilms applied in bioengineering systems. An adsorption of phenanthrene was used for analysis of the hydrophobicity of support fibers and bacterial cell surfaces within the biofilter of wastewater. The adsorption of phenanthrene was measured by synchronous fluorescence spectrometry. Cell surface hydrophobicity does not depend on the fixation procedure, pH of microbial suspension, and has no clear correlation with an adherence of the cells to hexadecane droplets. Notwithstanding high hydrophobicity of bacterial cells, the hydrophobicity of intact biofilm is determined by the hydrophobicity of the support fibers. New indexes were proposed to evaluate the reactor performance related with hydrophobic interactions within the biofilm. These indexes showed that significant share of hydrophobic sites within the nitrifying biofilm is protected from the hydrophobic interactions between the cells and environment.     

Population dynamics and in situ kinetics of nitrifying bacteria in autotrophic nitrifying biofilms as determined by real-time quantitative PCR

Population dynamics of ammonia-oxidizing bacteria (AOB) and uncultured Nitrospira-like nitrite-oxidizing bacteria (NOB) dominated in autotrophic nitrifying biofilms were determined by using real-time quantitative polymerase chain reaction (RTQ-PCR) and fluorescence in situ hybridization (FISH). Although two quantitative techniques gave the comparable results, the RTQ-PCR assay was easier and faster than the FISH technique for quantification of both nitrifying bacteria in dense microcolony-forming nitrifying biofilms. Using this RTQ-PCR assay, we could successfully determine the maximum specific growth rate (mu = 0.021/h) of uncultured Nitrospira-like NOB in the suspended enrichment culture. The population dynamics of nitrifying bacteria in the biofilm revealed that once they formed the biofilm, the both nitrifying bacteria grew slower than in planktonic cultures. We also calculated the spatial distributions of average specific growth rates of both nitrifying bacteria in the biofilm based on the concentration profiles of NH4+, NO2-, and O2, which were determined by microelectrodes, and the double-Monod model. This simple model estimation could explain the stratified spatial distribution of AOB and Nitrospira-like NOB in the biofilm. The combination of culture-independent molecular techniques and microelectrode measurements is a very powerful approach to analyze the in situ kinetics and ecophysiology of nitrifying bacteria including uncultured Nitrospira-like NOB in complex biofilm communities.     

Fate of 14C-labeled microbial products derived from nitrifying bacteria in autotrophic nitrifying biofilms

The cross-feeding of microbial products derived from 14C-labeled nitrifying bacteria to heterotrophic bacteria coexisting in an autotrophic nitrifying biofilm was quantitatively analyzed by using microautoradiography combined with fluorescence in situ hybridization (MAR-FISH). After only nitrifying bacteria were labeled with [14C]bicarbonate, biofilm samples were incubated with and without NH4+ as a sole energy source for 10 days. The transfer of 14C originally incorporated into nitrifying bacterial cells to heterotrophic bacteria was monitored with time by using MAR-FISH. The MAR-FISH analysis revealed that most phylogenetic groups of heterotrophic bacteria except the beta-Proteobacteria showed significant uptake of 14C-labeled microbial products. In particular, the members of the Chloroflexi were strongly MAR positive in the culture without NH4+ addition, in which nitrifying bacteria tended to decay. This indicated that the members of the Chloroflexi preferentially utilized microbial products derived from mainly biomass decay. On the other hand, the members of the Cytophaga-Flavobacterium cluster gradually utilized 14C-labeled products in the culture with NH4+ addition in which nitrifying bacteria grew. This result suggested that these bacteria preferentially utilized substrate utilization-associated products of nitrifying bacteria and/or secondary metabolites of 14C-labeled structural cell components. Our results clearly demonstrated that the coexisting heterotrophic bacteria efficiently degraded and utilized dead biomass and metabolites of nitrifying bacteria, which consequently prevented accumulation of organic waste products in the biofilm.     

Ecophysiological interaction between nitrifying bacteria and heterotrophic bacteria in autotrophic nitrifying biofilms as determined by microautoradiography-fluorescence in situ hybridization

Ecophysiological interactions between the community members (i.e., nitrifiers and heterotrophic bacteria) in a carbon-limited autotrophic nitrifying biofilm fed only NH(4)(+) as an energy source were investigated by using a full-cycle 16S rRNA approach followed by microautoradiography (MAR)-fluorescence in situ hybridization (FISH). Phylogenetic differentiation (identification) of heterotrophic bacteria was performed by 16S rRNA gene sequence analysis, and FISH probes were designed to determine the community structure and the spatial organization (i.e., niche differentiation) in the biofilm. FISH analysis showed that this autotrophic nitrifying biofilm was composed of 50% nitrifying bacteria (ammonia-oxidizing bacteria [AOB] and nitrite-oxidizing bacteria [NOB]) and 50% heterotrophic bacteria, and the distribution was as follows: members of the alpha subclass of the class Proteobacteria (alpha-Proteobacteria), 23%; gamma-Proteobacteria, 13%; green nonsulfur bacteria (GNSB), 9%; Cytophaga-Flavobacterium-Bacteroides (CFB) division, 2%; and unidentified (organisms that could not be hybridized with any probe except EUB338), 3%. These results indicated that a pair of nitrifiers (AOB and NOB) supported a heterotrophic bacterium via production of soluble microbial products (SMP). MAR-FISH revealed that the heterotrophic bacterial community was composed of bacteria that were phylogenetically and metabolically diverse and to some extent metabolically redundant, which ensured the stability of the ecosystem as a biofilm. alpha- and gamma-Proteobacteria dominated the utilization of [(14)C]acetic acid and (14)C-amino acids in this biofilm. Despite their low abundance (ca. 2%) in the biofilm community, members of the CFB cluster accounted for the largest fraction (ca. 64%) of the bacterial community consuming N-acetyl-D-[1-(14)C]glucosamine (NAG). The GNSB accounted for 9% of the (14)C-amino acid-consuming bacteria and 27% of the [(14)C]NAG-consuming bacteria but did not utilize [(14)C]acetic acid. Bacteria classified in the unidentified group accounted for 6% of the total heterotrophic bacteria and could utilize all organic substrates, including NAG. This showed that there was an efficient food web (carbon metabolism) in the autotrophic nitrifying biofilm community, which ensured maximum utilization of SMP produced by nitrifiers and prevented buildup of metabolites or waste materials of nitrifiers to significant levels.     

Retention of bacteria on a substratum surface with micro-patterned hydrophobicity

Bacteria adhere to almost any surface, despite continuing arguments about the importance of physico-chemical properties of substratum surfaces, such as hydrophobicity and charge in biofilm formation. Nevertheless, in vivo biofilm formation on teeth and also on voice prostheses in laryngectomized patients is less on hydrophobic than on hydrophilic surfaces. With the aid of micro-patterned surfaces consisting of 10-microm wide hydrophobic lines separated by 20-microm wide hydrophilic spacings, we demonstrate here, for the first time in one and the same experiment, that bacteria do not have a strong preference for adhesion to hydrophobic or hydrophilic surfaces. Upon challenging the adhering bacteria, after deposition in a parallel plate flow chamber, with a high detachment force, however, bacteria were easily wiped-off hydrophobic lines, most notably when these lines were oriented parallel to the direction of flow. Adhering bacteria detached slightly less from the hydrophilic spacings in between, but preferentially accumulated adhering on the hydrophilic regions close to the interface between the hydrophilic spacings and hydrophobic lines. It is concluded that substratum hydrophobicity is a major determinant of bacterial retention while it hardly influences bacterial adhesion.     

Alginate as an auxiliary bacterial membrane: binding of membrane-active peptides by polysaccharides.

The chronicity of Pseudomonas aeruginosa infections in cystic fibrosis (CF) patients is characterized by overproduction of the exopolysaccharide alginate, in which biofilm bacteria are embedded. Alginate apparently contributes to the antibiotic resistance of bacteria in this form by acting as a diffusion barrier to positively charged antimicrobial agents. We have been investigating cationic antimicrobial peptides (CAPs) (prototypic sequence: KKAAAXAAAAAXAAWAAXAAAKKKK-NH(2), where X is any of the 20 commonly occurring amino acids) that were originally designed as transmembrane mimetic peptides. Peptides of this group above a specific hydrophobicity threshold insert spontaneously into membranes and have antibacterial activity at micromolar concentrations. While investigating the molecular basis of biofilm resistance to peptides, we found that the anionic alginate polysaccharide induces conformational changes in the most hydrophobic of these peptides typically associated with insertion of such peptides into membrane environments [Chan et al., J. Biol. Chem. (2004) vol. 279, pp. 38749-38754]. Through a combination of experiments measuring release of the fluorescent dye calcein from phospholipid vesicles, peptide interactions with vesicles in the presence and absence of alginate, and affinity of peptides for alginate as a function of net peptide core hydrophobicity, we show here that alginate offers a microenvironment that provides a protective mechanism for the encased bacteria by both binding and promoting the self-association of the CAPs. The overall results indicate that hydrophilic alginate polymers contain a significant hydrophobic compartment, and behave as an 'auxiliary membrane' for bacteria, thus identifying a unique protective role for biofilm exopolysaccharide matrices.     

Fate of 14C-Labeled Microbial Products Derived from Nitrifying Bacteria in Autotrophic Nitrifying Biofilms

The cross-feeding of microbial products derived from ¹⁴C-labeled nitrifying bacteria to heterotrophic bacteria coexisting in an autotrophic nitrifying biofilm was quantitatively analyzed by using microautoradiography combined with fluorescence in situ hybridization (MAR-FISH). After only nitrifying bacteria were labeled with [¹⁴C]bicarbonate, biofilm samples were incubated with and without NH₄⁺ as a sole energy source for 10 days. The transfer of ¹⁴C originally incorporated into nitrifying bacterial cells to heterotrophic bacteria was monitored with time by using MAR-FISH. The MAR-FISH analysis revealed that most phylogenetic groups of heterotrophic bacteria except the β-Proteobacteria showed significant uptake of ¹⁴C-labeled microbial products. In particular, the members of the Chloroflexi were strongly MAR positive in the culture without NH₄⁺ addition, in which nitrifying bacteria tended to decay. This indicated that the members of the Chloroflexi preferentially utilized microbial products derived from mainly biomass decay. On the other hand, the members of the Cytophaga-Flavobacterium cluster gradually utilized ¹⁴C-labeled products in the culture with NH₄⁺ addition in which nitrifying bacteria grew. This result suggested that these bacteria preferentially utilized substrate utilization-associated products of nitrifying bacteria and/or secondary metabolites of ¹⁴C-labeled structural cell components. Our results clearly demonstrated that the coexisting heterotrophic bacteria efficiently degraded and utilized dead biomass and metabolites of nitrifying bacteria, which consequently prevented accumulation of organic waste products in the biofilm.     

Microbial community structure in autotrophic nitrifying granules characterized by experimental and simulation analyses

This study evaluates the community structure in nitrifying granules (average diameter of 1600 μm) produced in an aerobic reactor fed with ammonia as the sole energy source by a multivalent approach combining molecular techniques, microelectrode measurements and mathematical modelling. Fluorescence in situ hybridization revealed that ammonia-oxidizing bacteria dominated within the first 200 μm below the granule surface, nitrite-oxidizing bacteria a deeper layer between 200 and 300 μm, while heterotrophic bacteria were present in the core of the nitrifying granule. Presence of these groups also became evident from a 16S rRNA clone library. Microprofiles of NH₄⁺, NO₂^–, NO₃^– and O₂ concentrations measured with microelectrodes showed good agreement with the spatial organization of nitrifying bacteria. One- and two-dimensional numerical biofilm models were constructed to explain the observed granule development as a result of the multiple bacteria–substrate interactions. The interaction between nitrifying and heterotrophic bacteria was evaluated by assuming three types of heterotrophic bacterial growth on soluble microbial products from nitrifying bacteria. The models described well the bacterial distribution obtained by fluorescence in situ hybridization analysis, as well as the measured oxygen, nitrite, nitrate and ammonium concentration profiles. Results of this study are important because they show that a combination of simulation and experimental techniques can better explain the interaction between nitrifying bacteria and heterotrophic bacteria in the granules than individual approaches alone.     

The Role of Cell Hydrophobicity in the Formation of Aerobic Granules

Cell hydrophobicity is an important affinity force in cell self-immobilization and attachment processes. The role of cell hydrophobicity in the formation of aerobic granules has not been clear. Therefore, two series of experiments were conducted to investigate the role of cell hydrophobicity in the formation of aerobic heterotrophic and nitrifying granules in sequencing batch reactors, while the effects of shear strength, hydraulic selection pressure, and organic loading rate on the cell hydrophobicity were also studied. Results showed that the formations of heterotrophic and nitrifying granules were associated very closely with the cell hydrophobicity. The hydrophobicity of granular sludge was nearly twofold higher than that of conventional bioflocs. A high shear force or hydraulic selection pressure imposed on microorganisms resulted in a significant increase in the cell hydrophobicity, while the cell hydrophobicity seemed not to be sensitive to the changes in the organic loading rates in the range studied. In conclusion, the cell hydrophobicity could induce and further strengthen cell–cell interaction, and might be a main triggering force to initiate the granulation of heterotrophic and nitrifying bacteria. Received: 21 May 2002 / Accepted: 21 June 2002     

In situ PCR for visualizing distribution of a functional gene "amoA" in a biofilm regardless of activity

In this study, ammonia-oxidizing bacteria present in biofilms resulting from a nitrifying reactor were detected by both a conventional FISH technique and an original in situ PCR technique. Both techniques showed that ammonia-oxidizing bacteria were found near the surface of the biofilms. However, after the biofilm had been exposed to 2 weeks of ammonia starvation, ammonia-oxidizing bacteria present in the biofilm could not be detected by fluorescence in situ hybridization (FISH) because they did not have sufficient copies of rRNA. In contrast, ammonia-oxidizing bacteria could be detected by in situ PCR with strong signal. It was thus demonstrated that a cell possessing a specific functional gene is detectable by in situ PCR regardless of its activity.     

焦化废水工业处理装置和实验室装置硝化菌群的分子比较

反应器的群落结构分析有助于对工业装置的故障原因进行诊断。为了解决某焦化废水处理装置硝化功能低下的故障,构建了一套相似的实验室装置作为参照系统,该装置的硝化功能良好。通过工业装置和实验室装置好氧池生物膜16SrDNA克隆文库的比较,分析了它们之间硝化菌群的组成差异。实验室装置克隆文库的构成说明Nitrosomonas europaea-Nitrosoccus mobilis类群和Nitrospira属Ⅰ亚区系分别是该工艺条件下优势的氨氧化菌和亚硝酸氧化菌,但工业装置的克隆文库中却没有找到任何与硝化菌序列相近的克隆,这说明工业装置中硝化菌的多度较低。进一步使用Taqman荧光探针实时定量PCR测定了样品中Nitrospira属的多度,实验室装置中Nitrospira属16S rDNA的拷贝数达到3.4×106个/微克基因组DNA,而工业装置的测定值不到实验室装置的1/300。这些试验结果都表明工业装置好氧池微生物群落中缺少适当的硝化菌群是造成其硝化能力低下的重要原因。提高菌群中Nitrosomonas属和Nitrospira属的多度是解决工业装置硝化能力低下的关键。     

Development of a simultaneous partial nitrification and anaerobic ammonia oxidation process in a single reactor

Up-flow oxygen-controlled biofilm reactors equipped with a non-woven fabric support were used as a single reactor system for autotrophic nitrogen removal based on a combined partial nitrification and anaerobic ammonium oxidation (anammox) reaction. The up-flow biofilm reactors were initiated as either a partial nitrifying reactor or an anammox reactor, respectively, and simultaneous partial nitrification and anammox was established by careful control of the aeration rate. The combined partial nitrification and anammox reaction was successfully developed in both biofilm reactors without additional biomass inoculation. The reactor initiated as the anammox reactor gave a slightly higher and more stable mean nitrogen removal rate of 0.35 (± 0.19) kg-N m⁻³ d⁻¹ than the reactor initiated as the partial nitrifying reactor (0.23 (± 0.16) kg-N m⁻³ d⁻¹). FISH analysis revealed that the biofilm in the reactor started as the anammox reactor were composed of anammox bacteria located in inner anoxic layers that were surrounded by surface aerobic AOB layers, whereas AOB and anammox bacteria were mixed without a distinguishable niche in the biofilm in the reactor started as the partial nitrifying reactor. However, it was difficult to efficiently maintain the stable partial nitrification owing to inefficient aeration in the reactor, which is a key to development of the combined partial nitrification and anammox reaction in a single biofilm reactor.     

Effect of electrode surface properties on enhanced electron transfer activity in microbial fuel cells

The influence of electrode surface chemistry over biofilm growth was evaluated for photo‐bioelectrocatalytic fuel cell. A consortium of photosynthetic bacteria was grown onto different electrodes designed with polyethylenimine (PEI) and multiwall carbon nanotubes as hydrophilic and hydrophobic modifier, respectively. The designed electrodes were loaded with 0.08, 0.17, and 0.33 μg/cm² of PEI to change the hydrophilicity. However, 0.56, 0.72, and 0.83 mg/cm² of multiwall carbon nanotubes were used to alter the hydrophobicity of the electrodes. The surface chemistry of electrode and bio‐interaction was evaluated as a function of contact angle and biofilm formation. The results were compared with those obtained with a carbon paper electrode. The contact angle on the untreated electrode (carbon paper) was 118°, whereas for hydrophobic and hydrophilic electrodes, the maximum and minimum contact angles were 170° and 0°, respectively. Interestingly, the maximum biofilm growth (0.2275 g, wet basis) was observed on highly hydrophobic surface; however, the maximum electrochemical performance (246 mV) was shown by the most hydrophilic electrode surface. PEI‐based electrode with good biofilm formation showed comparatively higher electrogenic activity.     

Factors associated with adherence to and biofilm formation on polystyrene by Stenotrophomonas maltophilia: the role of cell surface hydrophobicity and motility

We tested 40 clinical Stenotrophomonas maltophilia strains to investigate the possible correlation between adherence to and formation of biofilm on polystyrene, and cell surface properties such as hydrophobicity and motility. Most of the strains were able to adhere and form biofilm, although striking differences were observed. Eleven (27.5%) of the strains were hydrophobic, with hydrophobicity greatly increasing as S. maltophilia attached to the substratum. A positive correlation was observed between hydrophobicity and levels of both adhesion and biofilm formation. Most of the isolates showed swimming and twitching motility. A highly significant negative correlation was observed between swimming motility and level of hydrophobicity. Hydrophobicity is thus a significant determinant of adhesion and biofilm formation on polystyrene surfaces in S. maltophilia.     

Rapid start-up of nitrifying MBBRs at low temperatures: nitrification,biofilm response and microbiome analysis

The moving bed biofilm reactor (MBBR), operated as a post carbon removal system, requires long start-up times in comparison to carbon removal systems due to slow growing autotrophic organisms. This study investigates the use of carriers seeded in a carbon rich treatment system prior to inoculation in a nitrifying MBBR system to promote the rapid development of nitrifying biofilm in an MBBR system at temperatures between 6 and 8 °C. Results show that nitrification was initiated by the carbon removal carriers after 22 h of operation. High throughput 16S-rDNA sequencing indicates that the sloughing period was a result of heterotrophic organism detachment and the recovery and stabilization period included a growth of Nitrosomonas and Nitrospira as the dominant ammonia oxidizing bacteria (AOB) and nitrite oxidizing bacteria (NOB) in the biofilm. Peripheral microorganisms such as Myxococcales, a rapid EPS producer, appear to have contributed to the recovery and stabilization of the biofilm.

     

Effect of temperature and free ammonia on nitrification and nitrite accumulation in landfill leachate and analysis of its nitrifying bacterial community by FISH

The cause of seasonal failure of a nitrifying municipal landfill leachate treatment plant utilizing a fixed biofilm was investigated by wastewater analyses and batch respirometric tests at every treatment stage. Nitrification of the leachate treatment plant was severely affected by the seasonal temperature variation. High free ammonia (NH3-N) inhibited not only nitrite oxidizing bacteria (NOB) but also ammonia oxidizing bacteria (AOB). In addition, high pH also increased free ammonia concentration to inhibit nitrifying activity especially when the NH4-N level was high. The effects of temperature and free ammonia of landfill leachate on nitrification and nitrite accumulation were investigated with a semi-pilot scale biofilm airlift reactor. Nitrification rate of landfill leachate increased with temperature when free ammonia in the reactor was below the inhibition level for nitrifiers. Leachate was completely nitrified up to a load of 1.5 kg NH4-N m(-3)d(-1) at 28 degrees C. The activity of NOB was inhibited by NH3-N resulting in accumulation of nitrite. NOB activity decreased more than 50% at 0.7 mg NH3-N L(-1). Fluorescence in situ hybridization (FISH) was carried out to analyze the population of AOB and NOB in the nitrite accumulating nitrifying biofilm. NOB were located close to AOB by forming small clusters. A significant fraction of AOB identified by probe Nso1225 specifically also hybridized with the Nitrosomonas specific probe Nsm156. The main NOB were Nitrobacter and Nitrospira which were present in almost equal amounts in the biofilm as identified by simultaneous hybridization with Nitrobacter specific probe Nit3 and Nitrospira specific probe Ntspa662.     

Determination of the cell surface hydrophobicity of oral bacteria using a modified hydrocarbon adherence method

Abstract Hydrophobic interactions between bacterial cell surfaces and colonisable substrates have been implicated in the mechanisms of bacterial adherence. However, current methods of assessing bacterial hydrophobicity as a function of adherence to liquid hydrocarbons (especially hexadecane) do not always produce accurate or reproducible results. Therefore, the present technique was developed using xylene. The hydrophobic surface properties of fresh and type strains of Bacteriodes gingivalis, Bacteriodes intermedius, Capnocytophaga spp., Streptococcus salivarius and Streptococcus sanguis suspended either in saliva ions buffer (SIB) or in saliva diluted in SIB were measured. In SIB the test strains were predominantly hydrophobic. The addition of saliva caused a significant reduction ( P < 0.05) in hydrophobicity compared to SIB alone, with 80% of the strains tested. Since oral bacteria will be suspended in saliva in vivo, it is concluded that bacteria in the oral cavity may be less hydrophobic than previous studies have suggested.     

Biofilm and capsule formation of the diatom Achnanthidium minutissimum are affected by a bacterium

Photoautotrophic biofilms play an important role in various aquatic habitats and are composed of prokaryotic and/or eukaryotic organisms embedded in extracellular polymeric substances (EPS). We have isolated diatoms as well as bacteria from freshwater biofilms to study organismal interactions between representative isolates. We found that bacteria have a strong impact on the biofilm formation of the pennate diatom Achnanthidium minutissimum. This alga produces extracellular capsules of insoluble EPS, mostly carbohydrates (CHO), only in the presence of bacteria (xenic culture). The EPS themselves also have a strong impact on the aggregation and attachment of the algae. In the absence of bacteria (axenic culture), A. minutissimum did not form capsules and the cells grew completely suspended. Fractionation and quantification of CHO revealed that the diatom in axenic culture produces large amounts of soluble CHO, whereas in the xenic culture mainly insoluble CHO were detected. For investigation of biofilm formation by A. minutissimum, a bioassay was established using a diatom satellite Bacteroidetes bacterium that had been shown to induce capsule formation of A. minutissimum. Interestingly, capsule and biofilm induction can be achieved by addition of bacterial spent medium, indicating that soluble hydrophobic molecules produced by the bacterium may mediate the diatom/bacteria interaction. With the designed bioassay, a reliable tool is now available to study the chemical interactions between diatoms and bacteria with consequences for biofilm formation.     

Adhesion and biofilm formation on polystyrene by drinking water-isolated bacteria

This study was performed in order to characterize the relationship between adhesion and biofilm formation abilities of drinking water-isolated bacteria (Acinetobacter calcoaceticus, Burkholderia cepacia, Methylobacterium sp., Mycobacterium mucogenicum, Sphingomonas capsulata and Staphylococcus sp.). Adhesion was assessed by two distinct methods: thermodynamic prediction of adhesion potential by quantifying hydrophobicity and the free energy of adhesion; and by microtiter plate assays. Biofilms were developed in microtiter plates for 24, 48 and 72 h. Polystyrene (PS) was used as adhesion substratum. The tested bacteria had negative surface charge and were hydrophilic. PS had negative surface charge and was hydrophobic. The free energy of adhesion between the bacteria and PS was > 0 mJ/m² (thermodynamic unfavorable adhesion). The thermodynamic approach was inappropriate for modelling adhesion of the tested drinking water bacteria, underestimating adhesion to PS. Only three (B. cepacia, Sph. capsulata and Staphylococcus sp.) of the six bacteria were non-adherent to PS. A. calcoaceticus, Methylobacterium sp. and M. mucogenicum were weakly adherent. This adhesion ability was correlated with the biofilm formation ability when comparing with the results of 24 h aged biofilms. Methylobacterium sp. and M. mucogenicum formed large biofilm amounts, regardless the biofilm age. Given time, all the bacteria formed biofilms; even those non-adherents produced large amounts of matured (72 h aged) biofilms. The overall results indicate that initial adhesion did not predict the ability of the tested drinking water-isolated bacteria to form a mature biofilm, suggesting that other events such as phenotypic and genetic switching during biofilm development and the production of extracellular polymeric substances (EPS), may play a significant role on biofilm formation and differentiation. This understanding of the relationship between adhesion and biofilm formation is important for the development of control strategies efficient in the early stages of biofilm development.     

Biofilm retention on surfaces with variable roughness and hydrophobicity

Biofilms on food processing equipment cause food spoilage and pose a hazard to consumers. The bacterial community on steel surfaces in a butcher's shop was characterized, and bacteria representative of this community enriched from minced pork were used to study biofilm retention. Stainless steel (SS) was compared to two novel nanostructured sol-gel coatings with differing hydrophobicity. Surfaces were characterized with respect to roughness, hydrophobicity, protein adsorption, biofilm retention, and community composition of the retained bacteria. Fewer bacteria were retained on the sol-gel coated surfaces compared to the rougher SS. However, the two sol-gel coatings did not differ in either protein adsorption, biofilm retention, or microbial community composition. When polished to a roughness similar to sol-gel, the SS was colonized by the same amount of bacteria as the sol-gel, but the bacterial community contained fewer Pseudomonas cells. In conclusion, biofilm retention was affected more by surface roughness than chemical composition under the condition described in this study.