Claire M. Jardine

Antimicrobial Resistance in Escherichia coli Isolates from Swine and Wild Small Mammals in the Proximity of Swine Farms and in Natural Environments in Ontario, Canada

ABSTRACT Wild animals not normally exposed to antimicrobial agents can acquire antimicrobial agent-resistant bacteria through contact with humans and domestic animals and through the environment. In this study we assessed the frequency of antimicrobial resistance in generic Escherichia coli isolates from wild small mammals (mice, voles, and shrews) and the effect of their habitat (farm or natural area) on antimicrobial resistance. Additionally, we compared the types and frequency of antimicrobial resistance in E. coli isolates from swine on the same farms from which wild small mammals were collected. Animals residing in the vicinity of farms were five times more likely to carry E. coli isolates with tetracycline resistance determinants than animals living in natural areas; resistance to tetracycline was also the most frequently observed resistance in isolates recovered from swine (83%). Our results suggest that E. coli isolates from wild small mammals living on farms have higher rates of resistance and are more frequently multiresistant than E. coli isolates from environments, such as natural areas, that are less impacted by human and agricultural activities. No Salmonella isolates were recovered from any of the wild small mammal feces. This study suggests that close proximity to food animal agriculture increases the likelihood that E. coli isolates from wild animals are resistant to some antimicrobials, possibly due to exposure to resistant E. coli isolates from livestock, to the resistance genes of these isolates, or to antimicrobials through contact with animal feed.

Rats, Cities, People, and Pathogens: A Systematic Review and Narrative Synthesis of Literature Regarding the Ecology of Rat-Associated Zoonoses in Urban Centers

Urban Norway and black rats (Rattus norvegicus and Rattus rattus) are the source of a number of pathogens responsible for significant human morbidity and mortality in cities around the world. These pathogens include zoonotic bacteria (Leptospira interrogans, Yersina pestis, Rickettsia typhi, Bartonella spp., Streptobacillus moniliformis), viruses (Seoul hantavirus), and parasites (Angiostrongylus cantonensis). A more complete understanding of the ecology of these pathogens in people and rats is critical for determining the public health risks associated with urban rats and for developing strategies to monitor and mitigate those risks. Although the ecology of rat-associated zoonoses is complex, due to the multiple ways in which rats, people, pathogens, vectors, and the environment may interact, common determinants of human disease can still be identified. This review summarizes the ecology of zoonoses associated with urban rats with a view to identifying similarities, critical differences, and avenues for further study.

Antimicrobial Resistance in Generic Escherichia coli Isolates from Wild Small Mammals Living in Swine Farm, Residential, Landfill, and Natural Environments in Southern Ontario, Canada

ABSTRACT To assess the impacts of different types of human activity on the development of resistant bacteria in the feces of wild small mammals, we compared the prevalences and patterns of antimicrobial resistance and resistance genes in generic Escherichia coli and Salmonella enterica isolates from fecal samples collected from wild small mammals living in four environments: swine farms, residential areas, landfills, and natural habitats. Resistance to antimicrobials was observed in E. coli isolates from animals in all environments: 25/52 (48%) animals trapped at swine farms, 6/69 (9%) animals trapped in residential areas, 3/20 (15%) animals trapped at landfills, and 1/22 (5%) animals trapped in natural habitats. Animals trapped on farms were significantly more likely to carry E. coli isolates with resistance to tetracycline, ampicillin, sulfisoxazole, and streptomycin than animals trapped in residential areas. The resistance genes sul2, aadA, and tet(A) were significantly more likely to be detected in E. coli isolates from animals trapped on farms than from those trapped in residential areas. Three S. enterica serotypes (Give, Typhimurium, and Newport) were recovered from the feces of 4/302 (1%) wild small mammals. All Salmonella isolates were pansusceptible. Our results show that swine farm origin is significantly associated with the presence of resistant bacteria and resistance genes in wild small mammals in southern Ontario, Canada. However, resistant fecal bacteria were found in small mammals living in all environments studied, indicating that environmental exposure to antimicrobials, antimicrobial residues, resistant bacteria, or resistance genes is widespread.

Ecology of Leptospira interrogans in Norway rats (Rattus norvegicus) in an inner-city neighborhood of Vancouver, Canada.

Background Leptospira interrogans is a bacterial zoonosis with a worldwide distribution for which rats (Rattus spp.) are the primary reservoir in urban settings. In order to assess, monitor, and mitigate the risk to humans, it is important to understand the ecology of this pathogen in rats. The objective of this study was to characterize the ecology of L. interrogans in Norway rats (Rattus norvegicus) in an impoverished inner-city neighborhood of Vancouver, Canada. Methodology/Principal Findings Trapping was performed in 43 city blocks, and one location within the adjacent port, over a 12 month period. Kidney samples were tested for the presence of L. interrogans using PCR and sequencing. A multivariable model was built to predict L. interrogans infection status in individual rats using season and morphometric data (e.g., weight, sex, maturity, condition, etc.) as independent variables. Spatial analysis was undertaken to identify clusters of high and low L. interrogans prevalence. The prevalence of L. interrogans varied remarkably among blocks (0–66.7%), and spatial clusters of both high and low L. interrogans prevalence were identified. In the final cluster-controlled model, characteristics associated with L. interrogans-infection in rats included weight (OR = 1.14, 95% CI = 1.07–1.20), increased internal fat (OR = 2.12, 95% CI = 1.06–4.25), and number of bite wounds (OR = 1.20, 95% CI = 0.96–1.49). Conclusions/Significance Because L. interrogans prevalence varied with weight, body fat, and bite wounds, this study suggests that social structure and interactions among rats may influence transmission. The prevalence and distribution of L. interrogans in rats was also highly variable even over a short geographic distance. These factors should be considered in future risk management efforts.

Geography, Deer, and Host Biodiversity Shape the Pattern of Lyme Disease Emergence in the Thousand Islands Archipelago of Ontario, Canada

In the Thousand Islands region of eastern Ontario, Canada, Lyme disease is emerging as a serious health risk. The factors that influence Lyme disease risk, as measured by the number of blacklegged tick (Ixodes scapularis) vectors infected with Borrelia burgdorferi, are complex and vary across eastern North America. Despite study sites in the Thousand Islands being in close geographic proximity, host communities differed and both the abundance of ticks and the prevalence of B. burgdorferi infection in them varied among sites. Using this archipelago in a natural experiment, we examined the relative importance of various biotic and abiotic factors, including air temperature, vegetation, and host communities on Lyme disease risk in this zone of recent invasion. Deer abundance and temperature at ground level were positively associated with tick abundance, whereas the number of ticks in the environment, the prevalence of B. burgdorferi infection, and the number of infected nymphs all decreased with increasing distance from the United States, the presumed source of this new endemic population of ticks. Higher species richness was associated with a lower number of infected nymphs. However, the relative abundance of Peromyscus leucopus was an important factor in modulating the effects of species richness such that high biodiversity did not always reduce the number of nymphs or the prevalence of B. burgdorferi infection. Our study is one of the first to consider the interaction between the relative abundance of small mammal hosts and species richness in the analysis of the effects of biodiversity on disease risk, providing validation for theoretical models showing both dilution and amplification effects. Insights into the B. burgdorferi transmission cycle in this zone of recent invasion will also help in devising management strategies as this important vector-borne disease expands its range in North America.

The Characteristics of Wild Rat (Rattus spp.) Populations from an Inner-City Neighborhood with a Focus on Factors Critical to the Understanding of Rat-Associated Zoonoses

Norway and black rats (Rattus norvegicus and Rattus rattus) are among the most ubiquitous urban wildlife species and are the source of a number of zoonotic diseases responsible for significant human morbidity and mortality in cities around the world. Rodent ecology is a primary determinant of the dynamics of zoonotic pathogens in rodent populations and the risk of pathogen transmission to people, yet many studies of rat-associated zoonoses do not account for the ecological characteristics of urban rat populations. This hinders the development of an in-depth understanding of the ecology of rat-associated zoonoses, limits comparability among studies, and can lead to erroneous conclusions. We conducted a year-long trapping-removal study to describe the ecological characteristics of urban rat populations in an inner-city neighborhood of Vancouver, Canada. The study focused on factors that might influence the ecology of zoonotic pathogens in these populations and/or our understanding of that ecology. We found that rat population density varied remarkably over short geographical distances, which could explain observed spatial distributions of rat-associated zoonoses and have implications for sampling and data analysis during research and surveillance. Season appeared to influence rat population composition even within the urban environment, which could cause temporal variation in pathogen prevalence. Body mass and bite wounds, which are often used in epidemiologic analyses as simple proxies for age and aggression, were shown to be more complex than previously thought. Finally, we found that factors associated with trapping can determine the size and composition of sampled rat population, and thus influence inferences made about the source population. These findings may help guide future studies of rats and rat-associated zoonoses.

Carriage of Clostridium difficile by Wild Urban Norway Rats (Rattus norvegicus) and Black Rats (Rattus rattus)

ABSTRACT Clostridium difficile is an important cause of enteric infections in humans. Recently, concerns have been raised regarding whether animals could be a source of C. difficile spores. Although colonization has been identified in a number of domestic species, the ability of commensal pests to serve as a reservoir for C. difficile has not been well investigated. The objective of this study was to determine whether urban rats (Rattus spp.) from Vancouver, Canada, carry C. difficile. Clostridium difficile was isolated from the colon contents of trapped rats and was characterized using ribotyping, toxinotyping, and toxin gene identification. Generalized linear mixed models and spatial analysis were used to characterize the ecology of C. difficile in rats. Clostridium difficile was isolated from 95 of 724 (13.1%) rats, although prevalence differed from 0% to 46.7% among city blocks. The odds of being C. difficile positive decreased with increasing weight (odds ratio [OR], 0.67; 95% confidence interval [CI], 0.53 to 0.87), suggesting that carriage is more common in younger animals. The strains isolated included 9 ribotypes that matched recognized international designations, 5 identified by our laboratory in previous studies, and 21 “novel” ribotypes. Some strains were clustered geographically; however, the majority were dispersed throughout the study area, supporting environmental sources of exposure and widespread environmental contamination with a variety of C. difficile strains. Given that urban rats are the source of a number of other pathogens responsible for human morbidity and mortality, the potential for rats to be a source of C. difficile for humans deserves further consideration.

Carriage of methicillin-resistant Staphylococcus aureus by wild urban Norway rats (Rattus norvegicus).

Methicillin-resistant Staphylococcus aureus (MRSA) is an important cause of multi-drug-resistant infections in people, particularly indigent populations. MRSA can be transmitted between people and domestic animals, but the potential for transmission between people and commensal pests, particularly rodents, had not been investigated. The objective of this study was to identify the presence and characterize the ecology of MRSA in rats (Rattus spp.) from in an impoverished, inner-city neighborhood. Oropharyngeal swabs were collected from rats trapped in 33 city blocks and one location within the adjacent port. Bacterial culture was performed and MRSA isolates were characterized using a variety of methods, including whole-genome sequencing (WGS). The ecology of MRSA in rats was described using phylogenetic analysis, geospatial analysis, and generalized linear mixed models. MRSA was identified 22 of 637 (3.5%) rats tested, although prevalence varied from 0 – 50% among blocks. Isolates belonged to 4 clusters according to WGS, with the largest cluster (n = 10) containing isolates that were genetically indistinguishable from community-acquired USA300 MRSA strains isolated from people within the study area. MRSA strains demonstrated both geographic clustering and dispersion. The odds of an individual rat carrying MRSA increased with increased body fat (OR = 2.53, 95% CI = 1.33 – 4.82), and in the winter (OR = 5.29, 95% CI = 1.04 – 26.85) and spring (OR = 5.50, 95% CI = 1.10 – 27.58) compared to the fall. The results show that urban rats carried the same MRSA lineages occurring in local human and/or animal populations, supporting recent transmission from external sources. MRSA carriage was influenced by season, most likely as a result of temporal variation in rat behavior and rat-human interactions.

An enhanced technique combining pre-enrichment and passive filtration increases the isolation efficiency of Campylobacter jejuni and Campylobacter coli from water and animal fecal samples.

Improved isolation techniques from environmental water and animal samples are vital to understanding Campylobacter epidemiology. In this study, the efficiency of selective enrichment in Bolton Broth (BB) followed by plating on charcoal cefoperazone deoxycholate agar (CCDA) (conventional method) was compared with an approach combining BB enrichment and passive filtration (membrane method) adapted from a method previously developed for testing of broiler meat, in the isolation of thermophilic campylobacters from surface water and animal fecal samples. The conventional method led to recoveries of Campylobacter from 36.7% of the water samples and 78.0% of the fecal samples and similar numbers, 38.3% and 76.0%, respectively, were obtained with the membrane method. To investigate the genetic diversity of Campylobacter jejuni and Campylobacter coli obtained by these two methods, isolates were analyzed using Comparative Genomic Fingerprinting, a high-resolution subtyping technique. The conventional and membrane methods yielded similar numbers of Campylobacter subtypes from water (25 and 28, respectively) and fecal (15 and 17, respectively) samples. Although there was no significant difference in recovery rates between the conventional and membrane methods, a significant improvement in isolation efficiency was obtained by using the membrane method, with a false-positive rate of 1.6% compared with 30.7% obtained using the conventional method. In conclusion, although the two methods are comparable in sensitivity, the membrane method had higher specificity, making it a cost-effective procedure for the enhanced isolation of C. jejuni and C. coli from water and animal fecal samples.

Antimicrobial Resistance in Escherichia coli Isolates from Raccoons (Procyon lotor) in Southern Ontario, Canada

ABSTRACT We conducted a cross-sectional study to determine the prevalence of antimicrobial resistance (AMR) in fecal Escherichia coli isolates from raccoons (Procyon lotor) living in Ontario, Canada. From June to October 2007, we trapped raccoons in three areas: one primarily urban site around Niagara, one primarily rural site north of Guelph, and one at the Toronto Zoo. In addition, we conducted a longitudinal study at the Toronto Zoo site to investigate the temporal dynamics of fecal E. coli and AMR in raccoons. Reduced susceptibility to ≥1 antimicrobial agent was detected in E. coli isolates from 19% of 16 raccoons at the urban site, 17% of 29 raccoons from the rural site, and 42% of 130 samples collected from 59 raccoons at the zoo site. Raccoons from the zoo site were significantly more likely to shed E. coli with reduced susceptibility to ≥1 antimicrobial agent than animals from the rural site (odds ratio [OR], 3.41; 95% confidence interval [CI], 1.17 to 12.09; P = 0.02). Resistance to expanded-spectrum cephalosporins (and the associated bla CMY-2 gene) was detected in two animals from the zoo site and one animal from the rural site. Serotyping and pulsed-field gel electrophoresis analysis show that raccoons on the zoo grounds harbor a diverse assemblage of E. coli, with rapid bacterial turnover within individuals over time. Our study indicates that raccoons may shed resistant bacteria of public health significance and that raccoons have the potential to disseminate these bacteria throughout their environment.