Access Type

Open Access Dissertation

Date of Award

January 2025

Degree Type

Dissertation

Degree Name

Ph.D.

Department

Nutrition and Food Science

First Advisor

Yifan Zhang

Abstract

The complexity of community risk factors and transmission routes has been a major challenge to the containment of community-acquired antibiotic resistance. Possible factors range from environmental contamination, antibiotic misuse, and human exposure via animals, food, the environment, and human-to-human transmission. What has been missing from previous research is the integration of findings in animal and environmental sectors with bacteria implicated in community infections. To gain a better understanding of antibiotic resistance acquired in the community, a one health approach is clearly needed. Specifically, we aim to investigate the prevalence of antibiotic-resistant Enterococcus in wildlife, their gene profiles in comparison with bacteria from community-acquired infections. A total of 140 fecal samples were collected from birds and rodents across 10 areas in Metro-Detroit. Culture based isolation, PCR identification and antimicrobial susceptibility testing were done. Enterococcus was recovered from 102 (72.8%) samples which further resulted in 270 isolates and 268 were analyzed. Approximately 28% (74/268) of isolates were E. faecalis, 26% (71/268) were E. faecium, while 46% (123/268) belonged to other species. Resistance to at least one antibiotic was found in 97% of isolates, with the top three resistance phenotypes being lincomycin (89.55%), quinupristin/dalfopristin (Q/D) (32.84%), and nitrofurantoin (17.54 %). Multidrug resistant (MDR) was shown by 17% of the isolates. The cadmium resistance gene cadA was detected in 28% of isolates altogether but more prevalent in E. faecalis (90.5%), suggesting species variation in cadA acquisition and calling for further research on plasmid profiling as cadA is often associated with plasmids. The data demonstrates urban wildlife as a significant reservoir of antibiotic-resistant Enterococcus. To compare wildlife resistance patterns to human samples, resistance was profiled in human isolates by analyzing 81 vancomycin resistant Enterococcus (VRE) from outpatients who had no extensive antibiotic or hospital exposure at Henry Ford Health System from primarily urine (78%) and blood samples (12%). A total of 17 E. faecalis and 19 E. faecium were randomly selected based on areas where wildlife samples were collected and tested for antibiotic susceptibility resulting in 100% resistance to vancomycin and ciprofloxacin, and over 90% to erythromycin, kanamycin, lincomycin, and tylosin tartrate. Daptomycin and nitrofurantoin resistance was only found in E. faecium samples while Q/D resistance was only found in E. faecalis. cadA gene screening revealed a prevalence of 74% (60/81), including E. faecium (78%) and E. faecalis (68%), contrasting wildlife data. This can be explained by the different selective pressure bacteria encounter in the environment versus during treatment for chronic diseases. Further research is warranted to explore cadmium as an alternative selective pressure for antibiotic resistance in Enterococcus species. The higher resistances in human samples shown to multiple antibiotics in our study indicates that there are more selective pressures for transmission of antibiotics in hospitals than in wildlife posing a significant public health challenge. Finally, to explore genomic landscape of antibiotic resistance and virulence, whole genome sequencing (WGS), using in-silco multi-locus sequence typing (MLST), resistance gene profiling using Qiagen microbial insight - antimicrobial resistance (QMI-AR) database, and virulence gene analysis using virulence factor database (VFDB) was performed on 26 isolates (20 wildlife, 6 human). Fourteen sequence types (ST) were identified (8 from E. faecium and 6 from E. faecalis), with human isolates clustering tightly while wildlife isolates spread out. Thirty-two resistance genes across glycopeptides (human-dominant), aminoglycosides, and macrolides-lincosamides-streptogramines (MLS) (wildlife dominant), fluoroquinolones, tetracycline, and trimethoprim were identified. Two wildlife samples, 106a and 123b carried vanX-A, vanS-C, and vanXY-C genes while 3 tetracycline resistance genes, tet(L), tet(M), and tet(U) were found across 5 human and 2 wildlife isolates. efmA which confers resistance to fluoroquinolones was found in 9 samples. Similarly, ErmB which confers resistance against Q/D was also found in isolates 1 wildlife and 4 human isolates. Twenty-nine different virulence genes were identified in human and wildlife isolates across 4 different categories namely, adherence, biofilm formation, exoenzyme, and immune modulation with wildlife isolates like 13a and 19b mirroring human strain 106656 and 123b carrying van resistance genes and multiple virulence gene categories making them highly virulent. This overlap suggests a common reservoir between wildlife and human and the transmission potential between the two. The presence of isolates carrying a range of antibiotic resistance genes and virulence factors indicate potential public health concern. To summarize, urban wildlife is a reservoir of antibiotic-resistant Enterococcus carrying a diversity of antibiotic resistance genes and virulence genes. Our findings highlight the need for extensive monitoring of environmental contamination and their public health implication. Continued research is also needed to explore the role of metal contamination in the urban environment as well as the transmission potential of resistant strains from wildlife.

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