Our results suggest that CPE, as well as other MDR pathogens, occur in the coastal waters of a popular public recreational beach offshore Netanya, Israel and nearby river estuaries. These CPE appear to be abundant and well-acclimated to freshwater and seawater, as they were easy to isolate and at least on one occasion, the same E. bugadensis was found twice, in distant sampling sites, one month apart. It is important to state that at the time of our sampling (June and July 2018) no considerable fluctuations in seawater quality were reported(44). Residents and tourists visiting popular recreation and leisure sites may be exposed to these MDROs.
The fact that isolates with highly similar genomes were isolated from both the riverine and marine environments, implies a mutual source, however, its origin is still unclear. Rivers are putative reservoirs and sources of MDROs, which were shown to be virulent(16, 45) and MDROs are less frequently encountered in patients drowning in seawater(46). The pollution sources of Poleg River include discharge of urban sewage(47, 48), authorized dumping of treated sewage water(49) and cattle herding as seen on site. One of the main sources of Alexander River in the Nablus River, running from Nablus city through Tul-Karem located in the West Bank (Figure 1). Other possible contamination sources include effluents from the adjacent WWTPs, a sea-turtles rescue center, an algae plant, an agricultural catchment basin and reservoir waters(30, 50). Becker et al (2013) reported that since 1995 several polluting sources were successfully treated, yet Wadi Zeimar, a major contamination source concentrating pollutants from Tul-Karem to Nablus River, remained(31). The polluted rivers are the likely sources of marine contamination. Alternatively, coastal water can migrate up to e few kilometers inland, as shown by the daily salinity profiles measured by Ruppin Estuarine and Coastal Observatory(50), potentially reaching the possible contamination sources. Despite the limited mixing of water masses, bacterial cross-contamination may be substantial, hence bidirectional contamination is feasible.
Gut microbes may be adapted to the aquatic environment, in which their survival rates are poorly understood. Yet, cultivation-based studies demonstrate that common gut bacteria, such as Enterobacteriaceae, are frequently detected in freshwater(8, 10, 11, 15, 51, 52) and seawater(5-7). These bacteria may be able to cope with different salinities because the human gut environment is characterized by spatial and temporal heterogeneity in osmolarity, based on the kinds of meals consumed(53). The fitness of microbes and their growth rates depends not only on osmolarity but also on taxon-specific physiology and additional external factors such as nutrient abundance, pH and oxygen levels(54). The three sequenced CP Eneterobacter spp. genomes encoded proteins that are involved in halotolerance, including the osmosensitive K+ channel histidine kinase KdpD, NhaA type Na+/H+ antiporter DNA-binding protein H-NS(55). These sequences were found in scaffolds that were >400 kbp in length and were defined as chromosomal by plasFlow. This indicates that at least some of the salt tolerance-related traits in these strains are not linked to plasmids as in other bacteria, and therefore not to blaIMI genes, which most likely are encoded on plasmids. Salinity has been recently shown to be the most important factor modulating the distribution patterns of antibiotic resistance genes in oceans and river beach soils(56), although other studies of aquatic ecosystems failed to show this (57). Thus, it is feasible that less-studied traits that mediate salinity tolerance may be linked to antibiotic resistance genes.
Most importantly, these aquatic isolates are only remotely associated with the local clinical epidemiology. CP Enterobacter spp., are uncommon in our hospital’s clinical settings: between August 2013 and February 2019, we identified 129 Enterobacter spp. out of 798 CRE rectal screening isolates (16%). Carbapenemase-producing Enterobacter spp. were detected in 65 isolates. blaIMI genes were found only in three of these CPE isolates, while blaKPC (50 isolates) and blaNDM (10 isolates) were more common. However, the IMI mechanism can often go undetected, because only the five major enzymes (KPC, NDM, VIM, OXA-48 and IMP) are routinely tested. The clinical implication is that Enterobacter spp. carrying an unidentified blaIMI could have been misidentified as non-CP CREs. Since patients carrying non-CP CRE isolates are not cohorted in Israeli hospitals as CPE carriers, such misidentifications increase the potential for hospital cross-infection and outbreaks.
OXA-48 has also not been frequently encountered during this period in our facility (19 isolates out of 798 CREs, mostly seen in E. coli spp). Nevertheless, IMI carbapenemases appear to be emerging in clinical practice(58) and as well as causing nosocomial outbreaks(59).