Rosen Boshnakov

Increasing antibiotic resistance poses a serious threat to human and animal health, as well as to the well-being of the environment. The present study aimed to quantify the total enterococci and those resistant to five classes of antibiotics in the water of an urban stretch of the Yantra River, Bulgaria. Phenotypic and genotypic antibiotic resistance patterns of primary selected antibiotic-resistant enterococci were assessed by the disk diffusion method and PCR assays. In the urban river stretch, the total number and relative proportion of antibiotic-resistant enterococci depended on the water sampling point. The dynamics of the population density of enterococci and their associated antibiotic resistance along the river stretch indicated anthropogenic impact on water quality. Riverine Enterococcus spp. isolates showed high levels of multidrug resistance to ampicillin, erythromycin, ciprofloxacin, tetracycline, and high-level gentamicin, as well as phenotypic patterns similar to those in clinical isolates and isolates from untreated wastewater. Waterborne enterococci are a reliable indicator of the prevalence of antibiotic resistance in the waters of the Yantra River, reflecting anthropogenic impact and potential risk to human health associated with their intended use.

Antibiotic resistance (ABR) is a significant threat to human and animal health, as well as to the environment. Antibiotic residues, antibiotic-resistant bacteria and antibiotic resistance genes are emerging contaminants of water resources. The present study aimed to assess the prevalence of ABR among waterborne enterococci in an anthropogenic-affected section of the Yantra River (Bulgaria). The susceptibility of 426 strains to 13 antibiotics (ABs) was tested by the disk diffusion method, and the genes encoding resistance by PCR analyses. A total of 39% of isolates were found to be antibiotic-resistant, with 9% mainly being multidrug-resistant to three AB classes. The most common resistance was to erythromycin (19%), tetracycline (18%) and ampicillin (14%), encoded by the ermB, tetM and blaTEM genes. A total of 3% of isolates were ciprofloxacin-resistant and only 1% was resistant to vancomycin or high-level gentamicin. All isolates were susceptible to teicoplanin and linezolid. Spatial variations in ABR levels were found, with the lowest abundance of antibiotic-resistant enterococci occurring in upstream river waters, away from urban areas, and the highest in urban areas. The spread of waterborne antibiotic-resistant enterococci highlights the need for water pollution management, monitoring and control to limit anthropogenic pressures through wastewater discharges and diffuse fecal pollution, and to ensure the ecological well-being of receiving waters.

River biofilms were studied using culture-dependent and molecular methods to determine the phe¬notypic antibiotic resistance (ABR) of attached bacteria and the presence of resistance-encoding genes. A comparative assessment of the prevalence of ABR in river waters and water-associated biofilms was made. Benthic biofilms included Enterobacteriaceae and enterococci with lower levels of ABR compared to aquatic bacteria, and no multiple antibiotic resistance was detected. However, the identification of genet¬ic determinants described in clinical isolates (such as blaAmpC, blaSHV, blaVIM) in benthic biofilms indi¬cated the prevalence of ABR in the aquatic environment and the resulting risk to human health and water resources. The data obtained indicate a need to deepen the knowledge of biofilm communities in running water and their contribution to the acquisition and spread of ABR.

Waste water treatment plants (WWTP) are considered as a hotspot for the acquisition and dissemination of antimicrobial resistance (AMR). The present study aimed to assess the AMR rate of the waste water microbiome in a selected WWTP and the treatment efficiency. Culture-dependent and PCR methods were used in the AMR study of raw and treated waste water (TWW) microbiomes. The population proportion of heterotrophic plate count (HPC) bacteria resistant to five antibiotic classes was assessed, as well as the AMR phenotype of a total of 238 Enterobacteriaceae and 259 Enterococcus spp. strains. Waste water treatment increased tetracycline- and ciprofloxacin-resistant bacteria by 67% and 61%, as well as the incidence of Enterobacteriaceae resistant to ciprofloxacin, co-trimoxazole, and cephalosporins. Multiple resistance increased, and 8.8% of TWW isolates exhibited an ESBL-producing phenotype, most often encoded by blaTEM and blaCTX-M genes. The most common resistance among Enterococcus spp. was to erythromycin and tetracycline, and despite the increased AMR rate among TWW isolates, only the increase in tetracycline resistance and the decrease in high-level gentamicin resistance were significant. All parameters analysed demonstrated limited removal of resistant HPC or faecal indicator bacteria in the studied WWTP and a positive selective effect towards some of them, most often to ciprofloxacin.