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Destruction and injury of escherichia coli under vacuum microwave : death kinetics and transcriptional responses Yaghmaee, Parastoo

Abstract

Rapid development in microwave applications in the home and industry along with the increase in the possibility of exposure to microwave radiation have raised concerns about the effect of microwaves on living cells. Although numerous studies have been conducted, microwave effects on living cells still are not fully understood. Some scientists believe that the effect is solely attributable to microwave heating while others suggested that additional effects, other than thermal, are required to explain various types of molecular formations and alterations in a target organism. The present work was designed to study the effect of 2450 MHz microwave radiation under vacuum (vacuum microwave or VM) on kinetic parameters and transcriptional response of mid-stationary Escherichia coli (ATCC 11775) cells and to search for possible non- thermal effects associated with VM . In addition, the E. coli transcriptome in late-log and mid-stationary phase of growth was studied. In a preliminary study, the lethal effect of microwave radiation on the microorganisms naturally occurring on parsley during dehydration under vacuum was investigated. Fresh parsley leaves were dried with air-drying (AD) and vacuum microwave drying (VMD) at the same final temperature. This study showed that parsley leaves treated with VMD had lower microbial populations than AD samples at comparable water activity. In addition VMD was more effective against yeast and mould than against total aerobic populations. Since higher reduction in microbial population of fresh parsley leaves occurred not only in a shorter time but also at a lower final temperature as a result of VMD compared to AD, it can be concluded that VM drying was an effective method of reducing the number of naturally occurring microorganisms in parsley. Death kinetics of E. coli in peptone water were determined in a continuous-flow vacuum system with a water bath or microwave as the heating source. Vacuum was used to control the boiling point of water and to maintain the bacterial suspensions at specified temperatures (49°C to 64°C). The z value in the water bath under vacuum was 9.0 °C whereas for VM treatments at 510W and 711W it was 6.0 °C and 5.9 °C respectively, suggesting that E. coli is more sensitive to temperature changes during microwave heating than conventional heat treatments. Based upon the Arrhenius calculation of the activation energy it is proposed that the mechanism of E. coli inactivation in VM treatment is different from the inactivation that occurs during conventional heat treatment. Thus the impact of temperature on E. coli destruction under vacuum was not the same when microwaves were the medium of heat transfer. Further, a molecular biology approach, DNA microarray technology, was used to investigate E. coli transcriptional response to sub-lethal VM and water bath treatment at 50°C for 3 minutes. The results showed that the number of E. coli genes that their expression altered through water bath treatment was higher than during VM treatment. VM treatment had a larger effect on genes related to membrane structure and membrane transport systems suggesting that microwave destruction may follow the dielectric cell-membrane rupture theory. In addition VM affected the expression of genes encode f or enzymes related to metabolism of carbohydrates, lipids and amino acids to a greater extent than the water bath treatment. Conversely the effect of conventional water bath treatment on ribosomal subunits was higher. Although both treatments were employed under vacuum and signs of anaerobic respiration would be expected, there was more evidence at the transcriptional level for the start of anaerobic respiration in water bath treated cells than in VM treated cells. In the present work, the focus of the kinetic and gene expression studies was on stationary phase cells, while other gene expression studies have mostly worked with cells at the exponential phase of growth. To close the loop, another study was conducted to investigate the changes at the transcription 1evel in E. coli cells between late-exponential and mid-stationary phase of growth. In mid-stationary phase, genes encoding for energy metabolism as well as amino acids and carbohydrate metabolism were down regulated. In addition csg genes, required for curli synthesis, were induced and 70.5% of genes involved in cell motility were down regulated or were not detected in mid-stationary cells indicating that in this stage cells may have been less mobile and had more tendency to clump or stick to surfaces. The transcription of hupA, hupB, hlpA, himA and himD genes previously reported to show up-regulation upon entry into stationary phase were down regulated in mid-stationary cells suggesting that the mechanisms involved in cell function are not only different between lag, log and stationary phase of growth but also may differ in early, mid and late stationary phases.

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