The Distinguishing Effects of Low‐Intensity Electromagnetic Radiation
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Letters in Applied Microbiology ISSN 0266-8254 ORIGINAL ARTICLE The distinguishing effects of low-intensity electromagnetic radiation of different extremely high frequencies on Enterococcus hirae: growth rate inhibition and scanning electron microscopy analysis K. Hovnanyan1, V. Kalantaryan2 and A. Trchounian3 1 Institute of Molecular Biology of National Academy of Sciences of Armenia, Yerevan, Armenia 2 Department of Radiophysics of High Frequences and Telecommunications, Faculty of Radiophysics, Yerevan State University, Yerevan, Armenia 3 Department of Biochemistry, Microbiology and Biotechnology, Faculty of Biology, Yerevan State University, Yerevan, Armenia Significance and Impact of the Study: A stronger inhibitory effect of low-intensity electromagnetic field on Enterococcus hirae ATCC 9790 bacterial growth rate was observed with 53 GHz vs 51Á8 GHz, regard- less of exposure duration. Scanning electron microscopy analysis showed that almost all irradiated cells in the population have spherical shapes similar to nonirradiated ones, but they have increased diame- ters in case of irradiated cells at 53 GHz, but not 51Á8 GHz. The results are novel, showing distinguishing effects of low-intensity electromagnetic field of different frequencies. They could be applied in treat- ment of food and different products in medicine and veterinary, where E. hirae plays an important role. Keywords Abstract action mechanisms, bacterial growth, electromagnetic radiation, electron A low-intensity electromagnetic field of extremely high frequency has microscopy, Enterococcus hirae. inhibitory and stimulatory effects on bacteria, including Enterococcus hirae.It was shown that the low-intensity (the incident power density of À Correspondence 0Á06 mW cm 2) electromagnetic field at the frequencies of 51Á8 GHz and Armen Trchounian, Department of Biochem- 53 GHz inhibited E. hirae ATCC 9790 bacterial growth rate; a stronger effect istry, Microbiology and Biotechnology, Faculty was observed with 53 GHz, regardless of exposure duration (0Á5h,1h or of Biology, Yerevan State University, 1 Manoukian Str., 0025 Yerevan, Armenia. 2 h). Scanning electron microscopy analysis of these effects has been done; the E-mail: [email protected] cells were of spherical shape. Electromagnetic field at 53 GHz, but not 51Á8 GHz, changed the cell size—the diameter was enlarged 1Á3 fold at 2017/0191: received 30 January 2017, revised 53 GHz. These results suggest the difference in mechanisms of action on 5 May 2017 and accepted 6 June 2017 bacteria for electromagnetic fields at 51Á8 GHz and 53 GHz. doi:10.1111/lam.12764 It has been previously shown that the low-intensity Introduction EMF at the frequencies of 51Á8 GHz and 53 GHz inhib- Electromagnetic field (EMF) of extremely high frequencies ited Enterococcus hirae growth (Ohanyan et al. 2008), pro- is an environmental stress factor, which has different stimu- longing lag growth phase duration and decreasing specific lative and depressive effects on living organisms, including growth rate during the log growth phase. Interestingly, bacteria (Soghomonyan et al. 2016). The interest to study the effects have increased with the EMF frequency from EMF effects is of significance because of developing electro- 45 GHz to 53 GHz, and 53 GHz had stronger effects magnetic technologies and widening of their applications (Ohanyan et al. 2008). Similar inhibitory effects have in telecommunications, environment control, as well as been reported with Escherichia coli (Trchounian et al. treatment of food and different medical and veterinary 2001; Torgomyan et al. 2013) and others, especially lactic products (Soghomonyan et al. 2016; Pan et al. 2017). acid bacteria (Soghomonyan and Trchounian 2013) at 220 Letters in Applied Microbiology 65, 220--225 © 2017 The Society for Applied Microbiology K. Hovnanyan et al. Electromagnetic field effects on E. hirae different frequencies (Soghomonyan et al. 2016). Note dependent on the exposure duration of EMF: the inhibi- that Enterococci belong to lactic acid bacteria (Fisher and tory effect was observed at 0Á5 h of exposure, the stron- Phillips 2009). In addition, the changes in E. coli cell gest effect was at 1 h of exposure and the decrease at 2 h morphology, showing increased cellular sizes, have been was only a little lower one than that at 1 h of exposure determined (Torgomyan et al. 2013). However, in con- (see Fig. 1). This confirms the results on inhibition of trast to E. coli, antibacterial effects with E. hirae were not growth rate and colony-forming activity reported before dependent on pH, EMF exposure duration and other for E. hirae for different growth conditions (Ohanyan factors (Ohanyan et al. 2008). et al. 2008). Moreover, there are probably some compen- The changes in water structure and properties (Golovl- satory cellular mechanisms which might prevent the eva et al. 1997; Torgomyan et al. 2011, 2013) and alter- increase in the EMF stress effects on bacteria (Soghomon- ations in E. hirae and E. coli cell plasma membrane yan et al. 2016). bioenergetic characteristics, ion transport and enzymatic Electron microscopy study of E. hirae irradiated by the properties (Trchounian et al. 2001; Torgomyan et al. EMF at 51Á8 GHz (Fig. 2) and 53 GHz (Fig. 3) showed 2012; Pham et al. 2016) can be considered as cellular that almost all cells in the population have spherical shapes mechanisms for the influence of the EMF on bacteria. similar to nonirradiated (control) bacteria; only several The change in sensitivity to different chemicals, including enterococci of oblong shape were seen. Changes in the aver- various antibiotics, is among the EMF effects (Dardalhon age diameter were observed during cell size measurements et al. 1981; Tadevosyan et al. 2008; Torgomyan et al. 2012, (Table 1). Such changes in cell size by irradiation of EMF 2013). But these effects were more strongly expressed in could be due to the intracellular hydration. Interestingly, E. hirae, than in E. coli (Torgomyan et al. 2012). It is destabilization of hydrogen bond structure in water near clear that cellular mechanisms of EMF effects should be the surface of the cell membrane and within the cell has studied further. These effects are of significance, since been suggested (Sinitsyn et al. 2000; Kuznetsov et al. 2006), E. hirae can be considered as a model organism for this can create conditions for cellular response. Gram-positive bacteria (Gaechter et al. 2012; Torgomyan Moreover, the marked increase in cell sizes for E. hirae et al. 2012); in addition, they are among commensal and grown till stationary growth phase (see Materials and pathogenic bacteria in nature (Vela et al. 2015), they can Methods) was observed by EMF at 53 GHz, but not be used as probiotics, applied in dairy production or, on 51Á8 GHz (see Table 1). This could be due to temporary the other hand, can be linked to different diseases in alterations in cell morphology by the EMF at 51Á8 GHz, human organism (Gilad et al. 1998; Fisher and Phillips as shown with E. coli (Torgomyan et al. 2013). It is likely 2009; Chan et al. 2012; El-Ghaish et al. 2015; Garcıa- to the mediated effects of EMF at these frequencies on Hernandez et al. 2016; Jain et al. 2016). Thus, structural E. coli, when the difference in the effects between and functional peculiarities of E. hirae, especially mem- 51Á8 GHz and 53 GHz is observed (Torgomyan et al. brane composition and properties, are the focus of this 2012, 2013). These effects could be due to the partial study. Moreover, the understanding of specific effects of absorbance of EMF energy of different value by the liquid EMF at 53 GHz, but not 51Á8 GHz is required; appropri- ate mechanisms would be suggested. This will allow the 0·9 clarification of the applications of EMF at different fre- quencies in food industry, biotechnology, medicine and ) 0·85 –1 veterinary fields, where E. hirae plays an important role. 0·8 The aim of this study was to provide scanning electron 0·75 microscopy analysis of E. hirae irradiated by EMF at 0·7 51Á8 GHz and 53 GHz, in addition to the effects on bac- terial cell growth. Special attention would be paid to dis- 0·65 tinguish between the effects of EMF at these two 0·6 frequencies. Spesific growth rate (h 0·55 0·5 Control 0·5 1·0 2·0 Results and discussion Exposure to EMF irradiation (h) Low-intensity EMF at the frequency of 51Á8 GHz and Figure 1 The changes in specific growth rate of Enterococcus hirae 53 GHz had inhibitory effects on E. hirae ATCC 9790 ATCC 9790 after irradiation by electromagnetic field at the frequen- growth. Indeed, specific growth rate during the log cies of 51Á8 GHz (■) and 53 GHz ( ) with different exposure dura- growth phase was decreased, and these effects were more tion. Control is nonirradiated cells. For details, see Materials and strongly expressed at 53 GHz (Fig. 1). The effects were methods. Letters in Applied Microbiology 65, 220--225 © 2017 The Society for Applied Microbiology 221 Electromagnetic field effects on E. hirae K. Hovnanyan et al. (a) (b) Figure 2 Scanning electron microscopy images of Enterococcus hirae ATCC 9790 cells in control (a) and after bacterial suspension irradiation by an electromagnetic field at the frequency of 51Á8 GHz with exposure of 0Á5 h (b), 1 h (c) and 2 h (d). Scale bar is shown. For details, see Materials (c) (d) and methods. [Colour figure can be viewed at wileyonlinelibrary.com] (water) medium, when 51Á8 GHz, but not 53 GHz is a 2015). This can trigger structural changes in the cell and resonant frequency for the water molecules (Sinitsyn et al. activity inducing different deformations. However, in 2000; Kuznetsov et al. 2006). Indeed, the absorption of spite of remarkable alterations in cell membrane-asso- water molecules at near ultraviolet region was shown to ciated activity, a resonant frequency for E. hirae, as well be more increased by the EMF at 51Á8 GHz, than at as for other bacteria, has not been yet established.