High levels of antibiotic resistance genes and opportunistic pathogenic bacteria indicators in urban wild bird feces

Highlights

• Wild urban bird feces contribute to the maintenance and dissemination of ARGs.

• OPs are abundant in wild bird feces, and some carry both ARGs and virulence genes.

• Bird fecal ARGs persist moderately and can propagate in impacted soil.

• Residual antibiotics ingestion by birds may partially contribute to their fecal ARGs.

Abstract

This study analyzed fresh feces from three common bird species that live in urban environments and interact with human communities. Antibiotic resistance genes (ARGs) encoding resistance to three major classes of antibiotics (i.e., tetracyclines, β-lactams, and sulfonamides) and the mobile genetic element integrase gene (intI1) were abundant (up to 10⁹, 10⁸, 10⁹, and 10¹⁰ copies/g dry feces for tetW, bla_TEM, sul1, and intI1, respectively), with relative concentrations surprisingly comparable to that in poultry and livestock that are occasionally fed antibiotics. Biomarkers for opportunistic pathogens were also abundant (up to 10⁷ copies/g dry feces) and the dominant isolates (i.e., Enterococcus spp. and Pseudomonas aeruginosa) harbored both ARGs and virulence genes. ARGs in bird feces followed first-order attenuation with half-lives ranging from 1.3 to 11.1 days in impacted soil. Although residual antibiotics were detected in the feces, no significant correlation was observed between fecal antibiotic concentrations and ARG relative abundance. Thus, other unaccounted factors likely contributed selective pressure for ARG maintenance. These findings highlight the contribution of wild urban bird feces to the maintenance and dissemination of ARGs, and the associated health risks.

Introduction

The global spread of antibiotic resistance poses a growing threat to public health (O’Neill, 2014) and requires improved understanding of ARG sources and vectors to mitigate the associated risks (Vikesland et al., 2017; Wu et al., 2018). The resistome carried by wildlife could be an important but underappreciated factor in ARG dissemination (Allen et al., 2010), and urbanization could amplify such risks as it fosters many overlapping habitats and frequent interactions between wildlife populations and humans (Magle et al., 2012).

Urban wild birds tend to have a relatively high population density (1.13–1.18 birds/resident) (Fuller et al., 2009) and opportunities to interact with human communities (Luniak, 2004; Magle et al., 2012) since urban settings generally have abundant food sources, low predator pressure, and a milder microclimate. Urban birds also have a high probability of exposure to water sources contaminated with antibiotics (Xu et al., 2016; Zhang et al., 2018), which may impact the abundance and diversity of ARGs within their gut and fecal microbiomes. Additionally, urban wild birds may serve as vectors for antibiotic resistant bacteria (ARB) derived from anthropogenic sources, as demonstrated by studies revealing clinically important ARB in bird feces (Ahlstrom et al., 2018; Hernandez et al., 2013). These bird-carried ARB and ARGs can be transferred to urban residents through swimming in feces-polluted waters, dermal contact with bird feces (or impacted soil) during outdoor recreational activities (Tsiodras et al., 2008), or inhalation of aerosolized fecal particles (Feddes et al., 1992). Therefore, it is important to investigate the abundance, diversity, and bacterial hosts of ARGs harbored by common urban wild birds.

Previous studies have conducted culture-based identification of ARB in bird feces (Dolejska and Literak, 2019; Wang et al., 2017) or analyzed ARGs harbored by wild birds near ARG hotspots (e.g., wastewater treatment plants (Marcelino et al., 2019) and ARG-polluted rivers (Wu et al., 2018)). However, there is a need to assess the relative abundance, diversity, and seasonal persistence of ARGs and their co-occurrence with virulence genes after transport and deposition by wild birds in densely populated environments to inform the associated risks.

In this study, fecal samples deposited by common urban wild birds (i.e., ducks, crows, and gulls) were collected from highly frequented sites in Houston metropolitan areas. ARGs and the mobile genetic element (MGE) integrase gene (intI1) were measured and compared with those found in poultry and livestock. These ARGs are commonly reported in the environment and were thus selected to facilitate comparison with other studies. Opportunistic pathogen levels were quantified by specific biomarker abundance, and their antibiotic resistance and potential virulence were evaluated by plate assays followed by PCR analyses. The natural attenuation of ARGs in impacted soil was then monitored to assess their persistence over different seasons. Possible selective pressure by residual antibiotics and their co-occurrence and correlation to ARG abundance were also considered.