Rapid, Noninvasive, and Nondestructive Method for Biofilm Imaging on Metallic Surfaces Using Active Thermography.

A simple, rapid, low-cost method was proposed for the imaging of Pseudomonas aeruginosa biofilms on metallic surfaces using an infrared camera. Stainless steel coupons were cooled to generate a thermal gradient in relation to biofilm for active thermography (AT). Both cooling and image acquisition times were optimized and the images obtained with AT were compared with those from scanning electron microscopy. A free software (Thermofilm) was developed for image processing and the results were compared with the software ImageJ, with good agreement (from 87.7 to 103.8%). Images of coupons treated with sanitizer (peracetic acid) were obtained to show the applicability of the proposed method for biofilm studies. All analytical steps could be performed in 3 min in a noncontact, nondestructive, low-cost, portable, and easy-to-use way.

Nanoemulsions containing Cymbopogon flexuosus essential oil: Development, characterization, stability study and evaluation of antimicrobial and antibiofilm activities.

The increase of microbial resistance generates the search for new substances with antimicrobial potential. The essential oil of Cymbopogon flexuosus (Lemongrass) stands out in the literature for its antimicrobial, insecticide and antioxidant properties, but it has high volatilization and low stability, and the nanoencapsulation of this oil could be an alternative to overcome these limitations. Thus, the objective of this study was to develop, for the first time, nanoemulsions containing the essential oil of C. flexuosus, through a method that does not use organic solvent and with temperature control to avoid the volatilization of the oil, characterize and evaluate of stability and the antimicrobial and antibiofilm activities of these nanoemulsions. Nanoemulsions presented adequate physicochemical characteristics (average size less than 200 nm, polydispersity index less than 0.3, negative zeta potential and acid pH) which were maintained during 90 days of storage, and the nanoencapsulation of the C. flexuosus oil enhanced its therapeutic efficacy against the microorganisms evaluated in this study compared to the free oil. These results are very promising because among the microorganisms that the nanoemulsion containing C. flexuosus was able to inhibit the formation of biofilm are the bacteria Pseudomonas aeruginosa and Staphylococcus aureus, which were recently listed by the World Health Organization as priority pathogens for development of new antibiotics.

Antimicrobial and antibiofilm activities of nanoemulsions containing Eucalyptus globulus oil against Pseudomonas aeruginosa and Candida spp.

Candida species are the main responsible microorganisms for causing fungal infections worldwide, and Candida albicans is most frequently associated with infectious processes. Pseudomonas aeruginosa is a gram-negative bacterium commonly found in immunocompromised patients. The infection persistence caused by these microorganisms is often related to antimicrobial resistance and biofilm formation. In this context, the objective of the present study was to prepare and characterize nanoemulsions containing Eucalyptus globulus oil and to verify its antimicrobial and antibiofilm activities against P. aeruginosa and Candida spp. The nanoemulsions had a size of approximately 76 nm, a polydispersity index of 0.22, a zeta potential of - 9,42 mV and a pH of approximately 5.0. The E. globulus oil was characterized by gas chromatography, being possible to observe its main components, such as 1-8-Cineol (75.8%), p- Cymene (7.5%), α-Pinene (7.4%) and Limonene (6.4%). The antimicrobial activity of the nanoemulsion was determined from the macrodilution tests and the cell viability curve, where the minimum fungicidal concentration of 0.7 mg/mL for C. albicans and 1.4 mg/mL for C. tropicalis and C. glabrata were obtained. However, the nanoemulsions did not present antimicrobial activity against P. aeruginosa, since it contains only 5% of the oil, being ineffective for this microorganism. The nanoencapsulated oil action against the formed biofilm was evaluated by atomic force microscopy and calcofluor staining, and the nanoemulsion was more efficient for two of the three Candida species when compared to free oil.

Antimicrobial activity of Melaleuca alternifolia nanoparticles in polymicrobial biofilm in situ.

Microbial biofilms represent a challenge in the treatment of infections, due to the low efficacy of the antimicrobials. This study evaluated the antimicrobial effect of nanoparticles of Melaleuca alternifolia (TTO) in dental biofilm. Thirty-eight volunteers used an oral device in situ in situ including four bovine enamel specimens for 07 days. From the fifth day four solutions were applied randomly for each specimen: Physiological Saline Solution (0.85% NaCl) (C+), Chlorhexidine 0.12% (CHX), M. alternifolia oil 0.3% (TTO), and a nanoparticle solution of 0.3% M. alternifolia oil (NPTTO). The nanoparticles of TTO were characterized for pH, IPD, medium size, zeta potential and Transmission Electron Microscopy. Antimicrobial activity was evaluated by viable microorganisms count and the structure of the biofilm by atomic force microscopy. The NPTTO presented pH 6.4, particle diameter of 197.9 ± 1 nm, polydispersion index of 0.242 ± 0.005, zeta potential of -7.12 mV and ±0:27 spherical shape. The C+ resulted in 100% of bacterial vitality, while CHX, TTO and NPTTO showed 34.2%, 51.4% and 25.8%, respectively. The AFM images showed biofilms with an average roughness of 350 nm for C+, 275 nm for CHX, 500 nm for TTO and 100 nm for NPTTO. The NPTTO demonstrated excellent antimicrobial activity in the biofilm formed in situ and will possibly be used in future for the treatment/prevention of oral biofilms.

Anti-biofilm activity of A22 ((S-3,4-dichlorobenzyl) isothiourea hydrochloride) against Pseudomonas aeruginosa: Influence on biofilm formation, motility and bioadhesion.

Bacterial biofilms are involved in various medical infections and for this reason it is of great importance understanding adhesion mechanisms of involved microorganisms is essential to develop new strategies of prevention and control. Different approaches have been used for preventing biofilm related infections in health care settings, such as use of surface coatings agents in medical implants. In this context, is necessary to explore new compounds with anti-biofilm activity. Thus, this study evaluated for the first time the action of A22 against biofilms of Pseudomonas aeruginosa PAO1 strain and multi-resistant clinical isolates on biotic and abiotic surfaces. A22 acts as inhibitor of the MreB protein of the bacterial cell wall, causing the rods to change shape to the coccoid form. In this work, A22 at subinhibitory concentrations was able to prevent biofilm formation, and atomic force microscopy images showed that A22 was highly effective in inhibiting adhesion on polyethylene surfaces. Pseudomonas aeruginosa PAO1 exhibited a strong ability to adhere to HeLa cells, and A22 inhibited the aggregation after 4 h of exposure. Swarming and twitching motilities were significantly altered by A22 at subinhibitory concentrations. Thus, by changing the shape of the bacterial cell, many properties can be affected, such as motility, surface adhesion and biofilm formation. This work presents A22 as a promising novel antibacterial or surface coating agent of medical materials.