Antibiotic Resistance in Enterobacteriaceae from Surface Waters in Urban Brazil Highlights the Risks of Poor Sanitation

Abstract

Surface waters are an unappreciated reservoir of antimicrobial resistance (AMR). Poor sanitation brings different species of environmental bacteria into contact, facilitating horizontal gene transfer. To investigate the role of surface waters as potential reservoirs of AMR, we studied the point prevalence of fecal contamination, AMR genes, and Enterobacteriaceae in an urban lake and rural river system in Northeast Brazil in comparison with a lake and sewer system in Northeast Ohio in the United States. Surface water samples were examined for evidence of human fecal contamination using microbial source tracking and screened for plasmid-mediated fluoroquinolone resistance and carbapenemase genes. Enterobacteriaceae were detected using selective agar followed by antimicrobial susceptibility testing and detection of AMR genes by microarray, and classified by repetitive sequence-based polymerase chain reaction and multilocus sequence typing. Concentrations of human fecal bacteria in the Brazilian urban lake and sewage in Northeast Ohio were similarly high. Filtered water samples from the Brazilian urban lake, however, showed the presence of bla _OXA-48, bla _KPC, bla _VIM-2, qnrS, and aac(6')-lb-cr, whereas only bla _VIM-2 was identified in raw sewage from Northeast Ohio. From the Brazilian urban lake, 85% of the Enterobacteriaceae (n = 40) cultured were resistant to at least one clinically important antibiotic, including ST131 Escherichia coli harboring the extended-spectrum beta-lactamase CTX-M. Although two isolates demonstrated polymyxin resistance, mcr-1/2 was not detected. Our findings indicate that surface waters in an urban Brazilian site can serve as an environmental reservoir of AMR and that improving wastewater treatment and sanitation generally may ameliorate AMR dissemination.

Figure 1.

Map of Dique do Cabrito identifying the number of copies of human Bacteroides spp. at each survey site. DCN indicates a site in the northern half of Dique do Cabrito and DCS indicates a southern site. This figure appears in color at www.ajtmh.org.

Figure 2.

Comparison between study sites of human Bacteroides quantities determined by microbial source tracking. Presents copies of DNA of human (HF8) cluster Bacteroides spp. identified by quantitative PCR in samples from filtered waters. Only one sample of raw sewage was obtained from a sewer system in Northeast Ohio (NEO). A sample from piped water from the Brazilian rural river system did not contain human (HF8) cluster Bacteroides spp. and is not represented.

Figure 3.

Resistance phenotypes of Escherichia coli and Klebsiella pneumoniae isolated from Dique do Cabrito. The relative contributions of each of the four resistance phenotypes listed are represented by a single bar per species and location. DCN indicates a site in the northern half of Dique do Cabrito and DCS indicates a southern site. ESBL = extended-spectrum beta-lactamase; TMP/SMX = trimethoprim/sulfamethoxazole.

Figure 4.

Dendrogram and band pattern generated by DiversiLab rep-PCR of Escherichia coli isolated from the study sites. Similarity ≥ 95% indicates clonally related strains. ESBL+ indicates a positive ESBL test by VITEK. “Sewer” designates isolates from raw sewage in Northeast Ohio, “DC” isolates from the urban lake in Brazil, and “Jenipapo” isolates from the rural river in Brazil. ESBL = extended-spectrum β-lactamases.