Cereus Biofilm Research Articles

Mechanism and application of camellia saponins against Bacillus cereus biofilm formation on different food contact surfaces

Bacillus cereus is a serious foodborne pathogen. In nature, it can form biofilms on the surface of food utensils and food processing equipment, making it difficult to remove. In this study, the model strains B. cereus ATCC 10987 and ATCC 14579 were selected as test strains to investigate the mechanism and inhibition of biofilm by camellia saponins (CS), in order to provide a new reference for the development of a natural antibiofilm. CS had a good inhibitory on biofilms of B. cereus ATCC 10987 and ATCC 14579, with minimal biofilm inhibitory concentration (MBIC) and minimal biofilm eradication concentration (MBEC) of 64 mg/mL and 128 mg/mL, respectively, and could reduce the exopolysaccharide and eDNA secretion of B. cereus by CLSM, thereby destroying the biofilm's structure and inhibiting the early adhesion and metabolism of individual bacteria within the biofilm. The qRT-PCR showed that CS can down-regulating the expression of nine biofilm formation related genes (spo0A, codY, rpoN, comER, sinR, plcR, papR, nprR and LuxS), thus directly or indirectly affect the biofilm formation. Combined with semi-quantitative detection of AI-2 signaling molecules, it is speculated that CS inhibits the B. cereus biofilm formation by inhibiting LuxS gene expression is to reduce the production of AI-2 and down-regulate the expressions of PlcR and PapR genes, thus inhibiting the quorum sensing mediated by LuxS/AI-2 and PlcR-PapR systems. Finally, the application of CS on stainless steel, ceramic and glass surfaces showed that the ceramic surface had greater biofilm inhibition. Moreover, CS showed a better antibiofilm effect as contact time increased.

Characterization of emetic Bacillus cereus biofilm formation and cereulide production in biofilm

Bacillus cereus is a well-known foodborne pathogen that can cause human diseases, including vomiting caused by emetic toxin, cereulide, requiring 105-108 cells per gram to cause the disease. The bacterial cells may be eliminated during processing, but cereulide can survive in most processing techniques due to its resistance to high temperatures, extreme pH and proteolytic enzymes. Herein, we reported dynamic processes of biofilm formation of four different types and cereulide production within the biofilm. Confocal laser scanning microscopy (CLSM) images revealed that biofilms of the four different types reach each stage at different time points. Among the extracellular polymeric substances (EPS) components of the four biofilms formed by the emetic B. cereus F4810/72 strain, proteins account for the majority. In addition, there are significant differences (p < 0.05) in the EPS components at the same stage among biofilms of different types. The time point at which cereulide was first detected in the four types of biofilms was 24 h. In the biofilm of B. cereus formed in ultra-high-temperature (UHT) milk, the first peak of cereulide appeared at 72 h. The cereulide content of the biofilms formed in BHI was mostly higher than that of the biofilms formed in UHT milk. This study contributes to a better understanding of food safety issues in the industry caused by biofilm and cereulide toxin produced by B. cereus.

Desalination of seawater using integrated microbial biofilm/cellulose acetate membrane and silver NPs/activated carbon nanocomposite in a continuous mode

The main objective of the present study was to desalinate seawater using Bacillus cereus gravel biofilm and cellulose acetate (CA) membranes with and without silver nanoparticles (AgNPs) as a potent and safe disinfectant for the treated water. Six desalination trials (I, II, III, IV, V and VI) were performed using the proposed biofilm/cellulose membrane. Results confirmed that Bacillus cereus gravel biofilm (microbial desalination) is the optimal system for desalination of seawater. It could achieve 45.0% RE (initial salinity: 44,478 mg/L), after only 3 h compared to the other tested treatments. It could also achieve 42, 42, 57, 43 and 59% RE for TDS, EC, TSS, COD and BOD, respectively. To overcome the problem of the residual salinity and reach complete elimination of salt content for potential reuse, multiple units of the proposed biofilm can be used in sequence. As a general conclusion, the Bacillus cereus biofilm system can be considered as remarkably efficient, feasible, rapid, clean, renewable, durable, environmentally friendly and easily applied technology compared to the very costly and complicated common desalination technologies. Up to our knowledge, this is the first time microbial biofilm was developed and used as an effective system for seawater desalination.

Apple cider vinegar exhibits promising antibiofilm activity against multidrug-resistant Bacillus cereus isolated from meat and their products.

Bacillus cereus (B. cereus) biofilm is grown not only on medical devices but also on different substrata and is considered a potential hazard in the food industry. Quorum sensing plays a serious role in the synthesis of biofilm with its surrounding extracellular matrix enabling irreversible connection of the bacteria. The goal of the current investigation was to ascertain the prevalence, patterns of antimicrobial resistance, and capacity for B. cereus biofilm formation in meat and meat products in Egypt. In all, 150 meat and meat product samples were used in this study. For additional bacteriological analysis, the samples were moved to the Bacteriology Laboratory. Thereafter, the antimicrobial, antiquorum sensing, and antibiofilm potential of apple cider vinegar (ACV) on B. cereus were evaluated. Out of 150 samples, 34 (22.67%) tested positive for B. cereus. According to tests for antimicrobial susceptibility, every B. cereus isolates tested positive for colistin and ampicillin but negative for ciprofloxacin and imipenem. The ability to form biofilms was present in all 12 multidrug-resistant B. cereus isolates (n = 12); of these, 6 (50%), 3 (25%), and 3 (25%) isolates were weak, moderate, and strong biofilm producers, respectively. It is noteworthy that the ACV demonstrated significant inhibitory effects on B. cereus isolates, with minimum inhibitory concentrations varying between 2 and 8 μg/ml. Furthermore, after exposing biofilm-producing B. cereus isolates to the minimum biofilm inhibitory concentrations 50 of 4 μg/ml, it demonstrated good antibiofilm activity (>50% reduction of biofilm formation). Strong biofilm producers had down-regulated biofilm genes (tasA and sipW) and their regulator (plcR) compared to the control group, according to reverse transcriptase quantitative polymerase chain reaction analysis. Our study is the first report, that spotlights the ACV activity against B. cereus biofilm and its consequence as a strong antibacterial and antibiofilm agent in the food industry and human health risk.

Role of Bacillus cereus biofilm formation behavior in virulence and pathogenic characteristics

AbstractThis study aimed to examine the potential link between Bacillus cereus biofilm formation, virulence, and pathogenicity. The biofilm formation abilities of nine B. cereus strains isolated from food and two reference strains (ATCC 10876 and ATCC 25621) were measured using a crystal violet assay. Among the tested strains, three strains (GIHE 617‐5, GIHE 86‐09, and GIHE 728‐17) and both reference strains were capable of biofilm formation. A positive correlation was obtained for higher cell surface hydrophobicity and increased biofilm formation. In contrast, HPLC analysis for elevated autoinducer‐2 (Al‐2) production revealed a negative impact on biofilm formation. PCR data indicated that all tested strains were capable of producing common B. cereus enterotoxins, including Hbl—A, C, and D, CytK, Nhe—B and C, EntFM, and BceT, but were negative for production of the emetic toxin cereulide and the pore‐forming toxin Hly II. Meanwhile, RT‐PCR data revealed a close correlation between high biofilm formation and the upregulation of several tested virulence genes for selected strains. However, elevated upregulation of virulence genes was not consistent in all of the higher biofilm‐forming strains. Cytotoxicity analysis revealed higher virulence characteristics compared to those of low biofilm‐forming strains.

Transcriptomic analysis of biofilm formation by Bacillus cereus under different carbon source conditions

Abstract Background Previous studies found differences in the utilization of different carbon sources during biofilm formation by Bacillus cereus. Illumina HiSeq high-throughput sequencing technology was used to investigate the changes in gene transcript levels in Bacillus cereus biofilm bacteria under different carbon source conditions. Results Compared with the control group, the number of differentially expressed genes in the glucose, maltose, lactose, and skim milk-supplemented groups was 351, 1136, 133, and 487, respectively. The results showed that the pathways involved in the differentially expressed genes were mainly distributed in glycolysis and pentose phosphate pathway, tricarboxylic acid cycle, amino acid metabolism, and fatty acid metabolism. The gene expression of enzymes related to acetoin synthesis from pyruvate was mostly upregulated in the glucose-supplemented group. The gene expression of enzymes related to pyruvate synthesis of branched-chain amino acids in the maltose-supplemented group was mostly upregulated. In the lactose-supplemented group, the gene expression of acetoin biosynthesis from pyruvate was upregulated. Pyruvate production through glycolysis pathway increased in the skim milk-supplemented group, but the metabolic capacity of the tricarboxylic acid cycle did not change significantly. Conclusion The content of pyruvate stored by Bacillus cereus biofilm bacteria through glycolysis or pentose phosphate pathway increased, but the carbon flux into the tricarboxylic acid cycle did not increase, which suggested that carbon fluxes in the extracellular polysaccharide synthesis pathway of the biofilm may be increased, resulting in increased biofilm biomass formation.

Bacillus cereus in the Artisanal Cheese Production Chain in Southwestern Mexico.

Bacillus cereus is associated with milk, dairy product, and dairy farm contamination. The aim of this study was to characterize strains of B. cereus in the small-scale artisanal cheese production chain in southwestern Mexico. 130 samples were collected. B. cereus isolation was performed on Mannitol Egg Yolk Polymyxin (MYP) agar. Genotyping, enterotoxigenic profile, and determination of genes involved in the formation of B. cereus biofilm were performed by PCR. An antimicrobial susceptibility test was made by broth microdilution assay. The phylogenetic analysis was performed by amplification and sequencing of 16s rRNA. B. cereus sensu lato was isolated and molecularly identified in 16 samples and B. cereus sensu stricto (B. cereus) was the most frequently isolated and identified species (81.25%). Of all the isolated B. cereus sensu lato strains, 93.75% presented at least one gene for some diarrheagenic toxins, 87.5% formed biofilms, and 18.75% were amylolytic. All B. cereus sensu lato strains were resistant to beta-lactams and folate inhibitors. A close phylogenetic relationship between isolates was found between the cheese isolates and the air isolates. Strains of B. cereus sensu lato were found in small-scale artisanal cheeses on a farm in southwestern Mexico.

Efektivitas Sanitizer Komersial Berbasiskan Asam Perasetat terhadap Biofilm Bacillus cereus

Bacillus cereus is known to have the ability to adhere and form biofilms on the surface of stainless steel that causes problems in the food industries. Bacterial biofilms generally can increase resistance to sanitizer treatment. This study aimed to evaluate the ability of peracetic acid-based commercial sanitizer to inactivate B. cereus biofilm on stainless steel (SS) surfaces. Biofilm of B. cereus ATCC 10876 was developed on SS surfaces and treated with 7 commercial peracetic acid-based sanitizers at their recommended dosages. Two sanitizers, i.e. B (peracetic acid and QAC) and F (peracetic acid and acidified water) showing the ability to inactivate B. cereus on solid media at concentration of 200, 400, and 800 ppm were further tested on biofilms with contact times of 1, 3, and 5 minutes. The 48 hours biofilms B. cereus contained 2.78-3.78 CFU/cm2. Both sanitizers B and F had significant effects in inactivating B. cereus biofilm. In general, sanitizer B could reduce more biofilm bacteria at any contact time than sanitizer F. Use of 200 ppm of sanitizer B or F 5 minutes could inactivate 3.04 log CFU/cm2 and 2.68 log CFU/cm2 biofilm, respectively. Exposure of B. cereus biofilm to peracetic acid-based sanitizer resulted in the damage of the extracellular matrix of the biofilms. This study showed that commercial sanitizers containing peracetic acid and quaternary ammonium compounds were effective in inactivating B. cereus biofilms.

SpoVG is an important regulator of sporulation and affects biofilm formation by regulating Spo0A transcription in Bacillus cereus 0\u20139

BackgroundBacillus cereus 0–9, a Gram-positive, endospore-forming bacterium isolated from healthy wheat roots in our previous research, is considered to be an effective biocontrol strain against several soil-borne plant diseases. SpoVG, a regulator that is broadly conserved among many Gram-positive bacteria, may help this organism coordinate environmental growth and virulence to survive. This study aimed to explore the multiple functions of SpoVG in B. cereus 0–9.MethodsThe gene knockout strains were constructed by homologous recombination, and the sporulation process of B. cereus 0–9 and its mutants were observed by fluorescence staining method. We further determined the spore yields and biofilm formation abilities of test strains. Transcriptional fusion strains were constructed by overlapping PCR technique, and the promoter activity of the target gene was detected by measuring its fluorescence intensity. The biofilm production and colonial morphology of B. cereus 0–9 and its mutants were determined to study the functions of the target genes, and the transcription level of the target gene was determined by qRT-PCR.ResultsAccording to observation of the sporulation process of B. cereus 0–9 in germination medium, SpoVG is crucial for regulating sporulation stage V of B. cereus 0–9, which is identical to that of Bacillus subtilis but differs from that of Bacillus anthracis. In addition, SpoVG could influence biofilm formation of B. cereus 0–9. The transcription levels of two genes closely related to biofilm-formation, sipW and calY, were downregulated in a ΔspoVG mutant. The role of SpoVG in regulating biofilm formation was further explored by deleting the genes abrB and sinR in the ΔspoVG mutant, respectively, generating the double mutant strains ΔspoVGΔabrB and ΔspoVGΔsinR. The phenotypes of these double mutants were congruent with those of the single abrB and sinR deletion strains, respectively, which showed increased biofilm formation. This indicated that spoVG was located upstream of abrB and sinR in the regulatory pathway of B. cereus biofilm formation. Further, the results of qRT-PCR and the luminescence intensity of transcriptional fusion strains indicated that spoVG gene deletion could inhibit the transcription of Spo0A.ConclusionsSpoVG, an important regulator in the sporulation of B. cereus, is located upstream of Spo0A and participates in regulation of biofilm formation of B. cereus 0–9 through regulating the transcription level of spo0A. Sporulation and biofilm formation are crucial mechanisms by which bacteria respond to adverse conditions. SpoVG is therefore an important regulator of Spo0A and is crucial for both sporulation and biofilm formation of B. cereus 0–9. This study provides a new insight into the regulatory mechanism of environmental adaptation in bacteria and a foundation for future studies on biofilm formation of B. cereus.

(+)-Terpinen-4-ol Inhibits Bacillus cereus Biofilm Formation by Upregulating the Interspecies Quorum Sensing Signals Diketopiperazines and Diffusing Signaling Factors.

Bacillus cereus is a Gram-positive endospore-forming foodborne pathogen that causes lethal food poisoning and significant economic losses, usually through biofilm- and endospore-induced recurrent cross- and postprocessing contamination. Due to the lack of critical inhibitory targets and control strategies, B. cereus biofilm contamination is a problem that urgently needs a solution. In this study, the antibacterial and antibiofilm activities of several natural potential bacterial quorum sensing (QS) interferers, a group of spice-originated monoterpenoids, were screened, and terpinen-4-ol effectively inhibited B. cereus growth and biofilm and spore germination with minimum growth inhibition and 50% biofilm inhibitory concentrations of 8 and 2 μmol/mL, respectively. FESEM/CLSM and phenotypic research illustrated that in addition to a decrease in the number of attached B. cereus cells, (+)-terpinen-4-ol also obviously reduced extracellular matrix synthesis, especially exopolysaccharides, and inhibited the swarming motility and protease activity of B. cereus. (+)-Terpinen-4-ol did not exert a significant effect on AI-2 signals in B. cereus. Accordingly, the B. cereus-produced interspecies QS signals diffusing signal factors (DSFs, C8-C15) and diketopiperazines (DKPs) were detected and identified here, which suppressed B. cereus biofilm formation in a concentration-dependent manner. (+)-Terpinen-4-ol significantly increased the levels of specific DSF and DKP signals in B. cereus and down-regulated the gene expression of some rpfB homologues in transcription level. Moreover, both DKPs and DSFs inhibited swarming motility and protease activity in B. cereus, while just the DSF signals 2-dodecenoic acid and 11-methyl-2-dodecenoic acid inhibited exopolysaccharide synthesis like (+)-terpinen-4-ol. In summary, B. cereus strains were found to produce nine DSF- and six DKP-type QS signaling molecules, which repressed B. cereus biofilm formation. (+)-Terpinen-4-ol was confirmed to be a promising antibacterial and antibiofilm agent against B. cereus upregulating DSFs and DKPs levels, and it could target the critical genes rpfB for DSFs turnover.

Effect of dry sanitizing methods on Bacillus cereus biofilm.

Bacillus cereus is a relevant foodborne pathogen and biofilm producer which can contaminate and persist in the processing environment of both high and low water activity foods. Because of this, it is crucial to understand better the resistance of this pathogen biofilm to different sanitation methods. The aim of this study was to evaluate the efficacy of dry sanitizing treatments against B. cereus biofilm formed on stainless steel (SS) and polypropylene (PP). Biofilm formation was held through the static method at 25 °C. After 4 days of incubation, coupons were exposed for up to 30 min to UV-C light, dry heat, gaseous ozone, 70% ethanol, and a commercial sanitizer. Sodium hypochlorite (200 mg/l) was also tested in two different pH values (7 and 11) for comparison purposes. In general, the surface material did not influence (p > 0.05) the performance of the treatments. From 10 min of exposure, 70% ethanol and the commercial product caused the lowest reductions on both surfaces. In addition, dry heat exhibited a poor performance on PP, with reductions < 1 log CFU/cm2. UV-C light on SS and PP and ozone on PP achieved reductions around 2 log CFU/cm2 after 30 min. The same level of reduction was obtained after 5 or 10 min using sodium hypochlorite (200 mg/l). Therefore, the results showed that dry sanitizing methods are not as effective as sodium hypochlorite against B. cereus biofilms. Further studies to evaluate the efficacy of the combination of dry methods are necessary.

Study of biofilm-influenced corrosion on X80 pipeline steel by a nitrate-reducing bacterium, Bacillus cereus, in artificial Beijing soil

The biofilm of Bacillus cereus on the surface of X80 pipeline steel was investigated from forming to shedding. Based on the observed biofilm morphology and pit analysis, it was found that B. cereus biofilm could stimulate X80 pipeline steel pitting corrosion, which was attributed to the nitrate reduction of bacteria beneath the biofilm. Electrochemical measurements and general corrosion rate results showed that B. cereus biofilm can better accelerate X80 pipeline steel corrosion compared to sterile solutions. Interestingly, the results also showed that thick biofilms had a slight tendency to inhibit the general corrosion process compared with its formation and exfoliation, which was confirmed by scanning Kelvin probe. The corrosion rate of X80 pipeline steel in artificial Beijing soil is closely related to the state of the biofilm, and nitrate reducing bacteria accelerates the occurrence of pits. The corresponding corrosion mechanisms are proposed.

Effects of yellow and red bell pepper (paprika) extracts on pathogenic microorganisms, cancerous cells and inhibition of survivin.

The present work examined the biomedical value of red and yellow bell pepper extracts (YME and RME) in terms of antioxidant, antibacterial and anticancer activities by in vitro and virtual studies. The yield of extract was 3.49% for RME and 2.92% for YME. The level of total phenols and total flavonoids significantly varied between the type of extracts, and it was higher in RME than that in YME. The extracts showed promising DPPH and ABTS free radical scavenging rates. The extracts showed an excellent antibacterial activity. The minimal inhibitory concentration (MIC) of RME was 0.20mgmL-1 for Bacillus cereus, 0.30mg mL-1 for Escherichia coli, 0.50mg mL-1 for Staphylococcus aureus and 0.60mg mL-1 and for Pseudomonas aeruginosa, while the MIC of YME was 0.40mg mL-1 for B. cereus, 0.40mg mL-1 for E. coli, 0.50mg mL-1 for S. aureus, and 0.60mg mL-1 for P. aeruginosa. TEM results demonstrated the cellular damage induced by RME in B. cereus biofilm. The RME did not show any cytotoxicity in normal NIH3T3 cells, but at 125μg mL-1 did a strong cytotoxicity in human lung cancer cell line A549 as evident by cytotoxicity assay, ROS and AO/EB staining. The virtual biological examination indicated that β-carotene from RME was a potential compound with higher docking energy against both targeted enzymes and proteins as - 14.30 for LpxC and - 15.59 for survivin. Therefore, it is recommended that RME is a better functional food with novel biomedical properties and it deserves further evaluation for its the novel molecules against multidrug resistant pathogens.

Biofilm formation capacity of Bacillus cereus on silicone, polyethylene terephthalate, Teflon, and aluminium food contact materials

Abstract Biofilms on food contact materials represent public health issues because they are resistant to cleaning and disinfection. This study aims to assess the Bacillus cereus biofilm formation capacity on silicone, polyethylene terephthalate, Teflon, and aluminium food contact materials. The biofilm biomass was analysed with the crystal violet assay method. We used the standard strain B. cereus CCM 2010, wild strain B. cereus 100 and spores of those two strains. The results show that both the vegetative form the bacteria and it spores form large amounts of biofilm on silicone, followed by polyethylene terephthalate, Teflon, and aluminium. More detailed analysis has shown that spores form more biomass on all materials in comparison to the vegetative form and that the standard strains form low levels of biofilm in contrast to the wild strains. Selecting proper material with the lowest biofilm formation potential can prevent or reduce food contamination and consequently increase food safety.

Relationship between antimicrobial activity and amphipathic structure of ginsenosides

Ginsenoside has a wide range of pharmacological activities, but little research has been conducted on its microbial pathogen inhibition and the relationship between its antimicrobial activity and saponin structure. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) or minimum fungicidal concentration (MFC) of a series of ginsenosides were used to investigate the antimicrobial activity of the ginsenosides. And the value of hydrophile–lipophile balance (HLB) was introduced as physicochemical factors to clarify the difference in bacteriostatic activity of saponins. The effects of active ginsenoside Rh2 on the Fusobacterium nucleatum and Bacillus cereus biofilm system were observed by calgary biofilm device (CBD) and fluorescence confocal microscopy (CLSM). The morphological characteristics were examined by using transmission electron microscope (TEM) to explore the cell damage. The antimicrobial effects were in the following order from high to low: saponins containing one glycosyl group > aglycones > two ligands > more than two ligands. Ginsenoside Rh2 and RK3 (hydrophile lipophilic balance value were both 1.15) showed high activity against the tested strains. Correlation analysis shows that HLB is associated with antimicrobial activity (p < 0.01). The antimicrobial activity increased with the HLB value of ginsenosides determined by changes in the length, the number, and the composition of sugar side chains. Specific polar ginsenosides cause bacterial death by destroying the membrane system. Based on their good broad-spectrum antibacterial activity, less polar ginsenosides have potential as bacteriostatic agents.

Benzalkonium bromide is effective in removing Bacillus cereus biofilm on stainless steel when combined with cleaning-in-place

Bacillus cereus is a common foodborne pathogen in dairy environment, which is hard to remove for its strong biofilm forming ability. In our study, Clean-in-place (CIP) regime and benzalkonium bromide (BK) were used in combination to enhance biofilm removal effectiveness. The CIP regime (1.5% NaOH at 65 °C for 10 min-water rinse-1% HNO3 at 65 °C for 10 min-water rinse) often used in dairy industry only achieved a 4.69 log reduction/cm2. A step of 5000 μg/mL BK following this CIP regime could effectively remove the biofilm cells on stainless steel, and biofilm cells were removed to below detection limit. The efficacy of BK in inhibiting Bacillus cereus was investigated. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of BK against Bacillus cereus were 6.25 μg/mL and 25 μg/mL. The antimicrobial activity of BK took effect by inducing intracellular proteins and electrolytes leakage. Moreover, the microstructure of extracellular matrix was broken under confocal microscopy (CLSM) and the amounts of proteins, polysaccharides and DNA in extracellular matrix reduced by 44.8%, 69.2% and 60.9% respectively after treatment of 5000 μg/mL BK. By detecting the zeta potential of biofilm extracellular matrix and contact angle assay, we found BK could bind to extracellular matrix with electrostatic interaction and increasing the hydrophilicity of biofilm, making biofilm removal easier. So, BK combined with CIP regime could be used as an effective strategy to remove Bacillus cereus biofilm.

Effect of Thymus ciliatus oil-based disinfectant solutions against bio-films formed by Bacillus cereus strains isolated from pasteurized-milk processing lines in Algeria

Bacillus cereus is a foodborne pathogen that often persists in dairy environments and is associated with food poisoning and spoilage. This spore-forming bacterium has a high propensity to develop biofilms onto dairy processing equipment and resists to chemical cleaning and disinfecting. This study deals with the in vitro application of thyme oil-based sanitizer solutions against biofilms formed by B. cereus genotypes which persist in pasteurized-milk processing lines. The effect of Thymus ciliatus essential oil on B. cereus planktonic cells and biofilms was assessed. The oil was tested alone and in combination with organic acids or industrial cleaning agents, in order to improve the removal of B. cereus recurrent genotypes. Minimal inhibitory concentrations of planktonic growth (MICs), biofilm formation (MBIC) and biofilm eradication (MBEC) of oil and organic acids were evaluated by microdilution assays. Thyme oil was more effective than organic acids against B. cereus planktonic growth, biofilm formation and established bio-films. High values of MICs were obtained for the three organic acids tested (3.5-4.5%) in comparison with those of essential oil (0.082-0.088%). The combination of oil with other antimicrobials as acetic acid, NaOH or HNO3 improves their effectiveness against B. cereus biofilms. These oil-based sanitizer solutions allow complete B. cereus biofilm eradication and should be an attractive candidate for the control and removal of biofilms in the dairy envi-ronment.

Effect of thyme essential oil against Bacillus cereus planktonic growth and biofilm formation.

The objective of this study was to determine the effect of thyme essential oil (TEO) on the planktonic growth and biofilm formation of Bacillus cereus (B. cereus). GC-MS analysis of TEO allowed the detection of 13 compounds, and the major constituents were p-cymene (29.7%), thymol (23.73%), γ-terpinene (16.21%), and 1,8-cineole (9.74%). TEO exhibited a minimum inhibitory concentration (MIC) value against planktonic B. cereus of 0.25mg/mL. The potent effect of TEO to inhibit the growth of planktonic B. cereus was due to cell membrane damage, as evidenced by reduced cell viability, protein changes, decreased intracellular ATP concentration, increased extracellular ATP concentration and cell membrane depolarization, and cellular morphological changes. In addition, TEO exerted a significant inhibitory effect on B. cereus biofilm formation, as confirmed by environmental scanning electron microscopic images. These findings suggested that TEO has the potential to be developed as a natural food additive to control foodborne contamination associated with B. cereus and its biofilm.

Extracellular electron transfer of Bacillus cereus biofilm and its effect on the corrosion behaviour of 316L stainless steel.

Here, a heterogeneous Bacillus cereus (B. cereus) biofilm on the surface of 316 L stainless steel (SS) was observed. With electrochemical measurement and surface analysis, it was found that B. cereus biofilm could inhibit SS pitting corrosion, attributing to the blocking effect of bacterial biofilm on extracellular electron transfer (EET). Differential pulse voltammetry (DPV) and cyclic voltammetry (CV) results also showed that B. cereus biofilm clearly impeded the EET. The proposed mechanism for the decreased corrosion rates of SS involves the interactions of extracellular polymeric substance (EPS) with SS and biofilm formation blocking electron transfer, preventing the passive layer from destroying. After biofilm formation following initial attachment of cells and EPS, electron transfer between SS and the cathodic depolarizer (oxygen) was hindered.

Impact of manganese and heme on biofilm formation of Bacillus cereus food isolates.

The objective of this study was to determine the impact of manganese (Mn2+) and heme on the biofilm formation characteristics of six B. cereus food isolates and two reference strains (ATCC 10987 and ATCC 14579). The data obtained from the crystal violet assay revealed that addition of a combination of Mn2+ and heme to BHI growth medium induced B. cereus biofilm formation. However, the induction of biofilm formation was strictly strain-dependent. In all of the induced strains, the impact of Mn2+ was greater than that of heme. The impact of these two molecules on the phenotypic characteristics related to biofilm formation, such as cell density, sporulation and swarming ability, was determined in a selected food isolate (GIHE 72–5). Addition of Mn2+ and heme to BHI significantly (p < 0.05) increased the number of cells, which was correlated with the results of crystal violet assays as well as scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) analyses. In addition, induced biofilms showed higher numbers of spores and greater resistance to benzalkonium chloride. The swarming ability of B. cereus planktonic cells was increased in the presence of Mn2+ and heme in BHI. The expression levels of a number of selected genes, which are involved in mobility and extracellular polymeric substances (EPS) formation in B. cereus, were positively correlated with biofilm formation in the presence of Mn2+ and heme in BHI. These results further confirming the role of these molecules in swarming mobility and making matrix components related to B. cereus biofilm formation. These data indicate that signaling molecules present in the food environment might substantially trigger B. cereus biofilm formation, which could pose a threat to the food industry.