## 4 Discussion  
The antimicrobial activity exerted by honey is one of its most well-established bioactivities described in many studies [11]. Honey affects the growth and survival of microorganisms via multiple mechanisms [14; 41]. Low pH, high sugar concentration, high osmolarity and the presence of antibacterial substances such as hydrogen peroxide and phytochemicals in honey provide a hostile environment for bacteria [42]. Consequently, honey is expected to contain a limited number and diversity of microorganisms [14]. Nevertheless, honey is not sterile since it is a natural product that is not processed [3; 5; 12; 28]. Several studies have reported isolated microorganisms from honey samples produced in various geographical locations [4; 7; 12; 18; 43]. Interestingly, the number of microorganisms in honey ranged widely, from 0 to several thousand colony forming units (CFUs) per gram, depending on the sample and its freshness [8; 9; 18]. [PERSON] et al. [18] reported low bacterial load in 33 out of 38 honey samples harvested in southern Italy. Similar results were reported by other researchers in Argentina [8], Morocco [15], Poland [12], Saudi Arabia [44], and Mexico [7]. [PERSON] et al. [27] studied the microbiological quality of honey from Argentina and their results were comparable to those reported in previous studies conducted in Argentina as well as other parts of the world [8; 12; 18; 28; 45]. Bacteria have also been isolated from stingless bee honey [4; 23; 42]. Moreover, there are few studies that have described the microbial communities of honey using culture-independent methods like next generation sequencing (NGS) [17; 46; 47; 48]. Honey bacteria exerting antibacterial activity have been successfully isolated by several research groups [6; 12; 49; 50; 51; 52]. There are also reports on the antibacterial activity exerted by various bacterial isolates from stingless bee (_Hetetortigona itama_) honey [34; 42]. [PERSON] et al. [28] screened more than two thousand bacterial strains isolated from six US domestic honey and two manuka honeys against seven bacteria and one mold but identification of bacterial isolates was not conducted. Recently, [PERSON] et al. [12] investigated the antimicrobial potential of 163 bacteria isolated from honey samples produced in Poland. A number of studies have demonstrated high antibacterial activity of Greek honeys against major nosocomial and foodborne pathogens [1; 13; 35; 53]. However, this is the first study aiming to isolate and characterize bacteria from diverse types of Greek honeys exhibiting antibacterial potential. In total, 2014 bacterial isolates were screened against five important human and foodborne pathogens (_S. aureus, P. aeruginosa, S._ Typhimurium, _A. baumannii,_ and _C. freundii_). Three of them belong to the \"ESKAPE\" pathogens group (_E. faecium, S. aureus, K. pneumoniae, A. buumannii, P. aeruginosa_, and _Enterobacter_ spp.). The World Health Organization (WHO) included ESKAPE in a list of pathogens for which new antimicrobial compounds are urgently needed. _A. baumannii_ and _P. aeruginosa_ are listed in the critical priority list of pathogens, whereas _S. aureus_ (MRSA) is in the high priority group [54; 55; 56]. Furthermore, multidrug-resistant _C. freundii_ is of major concern, often implicated in nosocomial infections [57; 58]. This is the first study regarding the antibacterial activity of honey bacterial isolates against _C. freundii_ and _A. baumannii._  
Moreover, in this study a higher number of bacterial isolates (2014) was screened against five pathogens, compared to relevant studies [4; 42; 51; 52]. The most susceptible pathogen was _S. aureus_, followed by _C. freundii, P. aeruginosa_, and _A. baumannii._ In contrast, _S._ Typhimurium was the least affected tested pathogen. A possible explanation might be that Gram-negative bacteria possess an outer membrane which often contributes to resistance to a wide range of antibacterial factors (for instance, loss of porins located in outer membrane). In agreement with our study, [PERSON] et al.