The present study investigated the effects of microwave (MW) radiation applied under a sublethal temperature on Escherichia coli. The experiments were conducted at a frequency of 18 GHz and at a temperature below 40°C to avoid the thermal degradation of bacterial cells during exposure. The absorbed power was calculated to be 1,500 kW/m 3 , and the electric field was determined to be 300 V/m. Both values were theoretically confirmed using CST Microwave Studio 3D Electromagnetic Simulation Software. As a negative control, E. coli cells were also thermally heated to temperatures up to 40°C using Peltier plate heating. Scanning electron microscopy (SEM) analysis performed immediately after MW exposure revealed that the E. coli cells exhibited a cell morphology significantly different from that of the negative controls. This MW effect, however, appeared to be temporary, as following a further 10-min elapsed period, the cell morphology appeared to revert to a state that was identical to that of the untreated controls. Confocal laser scanning microscopy (CLSM) revealed that fluorescein isothiocyanate (FITC)-conjugated dextran (150 kDa) was taken up by the MW-treated cells, suggesting that pores had formed within the cell membrane. Cell viability experiments revealed that the MW treatment was not bactericidal, since 88% of the cells were recovered after radiation. It is proposed that one of the effects of exposing E. coli cells to MW radiation under sublethal temperature conditions is that the cell surface undergoes a modification that is electrokinetic in nature, resulting in a reversible MW-induced poration of the cell membrane.The effects of MW radiation on microorganisms have been studied and debated for more than half a century (3,4,10,12,17,20,28,29,35). The nature of the debate surrounding this interaction has often referred to the existence of so-called specific microwave (MW) effects that are nonthermal in nature (4,10,13,17,20,28,29). Much has been published supporting the notion that a range of specific MW effects exist and can be identified in terms of their manifestations on cell physiology (2,4,10,13,27,28). For example, Dreyfuss and Chipley examined the effects of MW radiation (2.45 GHz) at sublethal temperatures on the metabolic activities of a range of enzymes expressed by the bacterium Staphylococcus aureus (10). These results suggested that MW radiation affected S. aureus cells in a way that could not have been explained solely by thermaleffect theories. It has also been found that Burkholderia cepacia bacteria could be wholly inactivated using MW radiation at sublethal temperatures at a frequency of 20 GHz (2). Samarketu et al. (25) examined the effects of MW radiation at a frequency of 9.575 GHz on the physiological behavior of Cyanobacterium dolium (Anabaena dolium). The authors suggested that MW radiation nonthermally induced different biological effects by changing the protein structures by differentially partitioning the ions and altering the rates and/or directions of biochemical reactions (25)...

The effect of electromagnetic field (EMF) exposures at the microwave (MW) frequency of 18 GHz, on four cocci, Planococcus maritimus KMM 3738, Staphylococcus aureus CIP 65.8T, S. aureus ATCC 25923 and S. epidermidis ATCC 14990T, was investigated. We demonstrate that exposing the bacteria to an EMF induced permeability in the bacterial membranes of all strains studied, as confirmed directly by transmission electron microscopy (TEM), and indirectly via the propidium iodide assay and the uptake of silica nanospheres. The cells remained permeable for at least nine minutes after EMF exposure. It was shown that all strains internalized 23.5 nm nanospheres, whereas the internalization of the 46.3 nm nanospheres differed amongst the bacterial strains (S. epidermidis ATCC 14990T~ 0%; Staphylococcus aureus CIP 65.8T S. aureus ATCC 25923, ~40%; Planococcus maritimus KMM 3738, ~80%). Cell viability experiments indicated that up to 84% of the cells exposed to the EMF remained viable. The morphology of the bacterial cells was not altered, as inferred from the scanning electron micrographs, however traces of leaked cytosolic fluids from the EMF exposed cells could be detected. EMF-induced permeabilization may represent an innovative, alternative cell permeability technique for applications in biomedical engineering, cell drug delivery and gene therapy.

The aim of the present review was to evaluate the literature suggesting that consideration be given to the existence of specific microwave (MW) effects on prokaryotic microorganisms; that is, effects on organisms that cannot be explained by virtue of temperature increases alone. This review considered a range of the reported effects on cellular components; including membranes, proteins, enzyme activity as well as cell death. It is concluded that the attribution of such effects to non-thermal mechanisms is not justified due to poor control protocols and because of the possibility that an unmeasurable thermal force, relating to instantaneous temperature (T (i)) that occurs during MW processing, has not been taken into account. However, due to this lack of control over T (i), it also follows that it cannot be concluded that these effects are not 'non-thermal'. Due to this ambiguity, it is proposed that internal 'micro'-thermal effects may occur that are specific to MW radiation, given its inherent unusual energy deposition patterning.

The present study developed and verified a 'cold' microwave (MW) treatment that could lead to the inactivation of two common pathogenic species of bacteria, Escherichia coli and Staphylococcus aureus, in raw meats. A number of experimental conditions were designed and tested to maximise MW exposure without overheating the samples. The non-thermal effect was maximised by multiple exposure to attain efficient MW threshold intensities. It was shown that at sub-lethal temperatures repeated exposure using high frequency MW radiation was significantly more effective in decontaminating bacteria in raw meats compared to a single exposure. It was concluded that non thermal inactivation of pathogenic bacteria in raw meats could be achieved at defined conditions using high frequency MW radiation.

Ivanova, E. (2009). A new sterilization technique of bovine pericardial biomaterial using microwave radiation. Tissue Engineering Part C methods, 15 (3), 445-454. A new sterilization technique of bovine pericardial biomaterial using microwave radiation AbstractBioprosthetic valves created from chemically treated natural tissues such as bovine pericardial biomaterial are used as heart valve scaffolds. Methods currently available for sterilization of biomaterial for transplantation include the application of gamma radiation and chemical sterilants. These techniques, however, can be problematic because they can be expensive and lead to a reduction in tissue integrity. Therefore, improved techniques are needed that are cost effective and do not disrupt the physical properties, functionality, and lifespan of the valvular leaflets. This study examined a novel technique using nonthermalmicrowave radiation that could lead to the inactivation of bacteria in bovine pericardial biomaterial without compromising valve durability. Two common pathogenic species of bacteria, Escherichia coli and Staphylococcus aureus, were used as test microorganisms. Optimized microwave parameters were used to determine whether inactivation of pathogenic bacteria from bovine pericardium could be achieved. In addition, the effect of microwave sterilization on tissue integrity was examined. The mechanical properties (assessed using dynamic mechanical analysis) and tensile strength testing (using a Universal Tensile Tester) as well as thermal analysis (using thermogravimetric analysis and differential scanning calorimetry) indicated that microwave sterilization did not compromise the functionality of bovine pericardial biomaterial. Scanning electron microscopy imaging and cytotoxicity testing also confirmed that the structure and biocompatibility of transplant biomaterial remained unaltered after the sterilization process. Results from the application of this newmicrowave (MW) sterilization technique to bovine pericardium showed that nearcomplete inactivation of the contaminant bacteria was achieved. It is concluded that nonthermal inactivation of pathogenic bacteria from bovine pericardial biomaterial could be achieved using microwave radiation. Disciplines

The effect of electromagnetic field (EMF) exposures at the microwave (MW) frequency of 18 GHz, on four cocci, Planococcus maritimus KMM 3738, Staphylococcus aureus CIP 65.8T, S. aureus ATCC 25923 and S. epidermidis ATCC 14990T, was investigated. We demonstrate that exposing the bacteria to an EMF induced permeability in the bacterial membranes of all strains studied, as confirmed directly by transmission electron microscopy (TEM), and indirectly via the propidium iodide assay and the uptake of silica nanospheres. The cells remained permeable for at least nine minutes after EMF exposure. It was shown that all strains internalized 23.5 nm nanospheres, whereas the internalization of the 46.3 nm nanospheres differed amongst the bacterial strains (S. epidermidis ATCC 14990T~ 0%; Staphylococcus aureus CIP 65.8TS. aureus ATCC 25923, ~40%; Planococcus maritimus KMM 3738, ~80%). Cell viability experiments indicated that up to 84% of the cells exposed to the EMF remained viable. The morphology of the bacterial cells was not altered, as inferred from the scanning electron micrographs, however traces of leaked cytosolic fluids from the EMF exposed cells could be detected. EMF-induced permeabilization may represent an innovative, alternative cell permeability technique for applications in biomedical engineering, cell drug delivery and gene therapy.