Virulence genes, antibiotic resistance and integrons in Escherichia coli strains isolated from synanthropic birds from Spain.

The aim of this study was to determine the presence of virulence genes and antibiotic resistance profiles in 164 Escherichia coli strains isolated from birds (feral pigeons, hybrid ducks, house sparrows and spotless starlings) inhabiting urban and rural environments. A total of eight atypical enteropathogenic E. coli strains were identified: one in a house sparrow, four in feral pigeons and three in spotless starlings. Antibiotic resistance was present in 32.9% (54) of E. coli strains. The dominant type of resistance was to tetracycline (21.3%), ampicillin (19.5%) and sulfamethoxazole (18.9%). Five isolates had class 1 integrons containing gene cassettes encoding for dihydrofolate reductase A (dfrA) and aminoglycoside adenyltransferase A (aadA), one in a feral pigeon and four in spotless starlings. To our knowledge, the present study constitutes the first detection of virulence genes from E. coli in spotless starlings and house sparrows, and is also the first identification worldwide of integrons containing antibiotic resistance gene cassettes in E. coli strains from spotless starlings and pigeons.

Connectivity at the human-wildlife interface: starling movements relate to carriage of E. coli.

Synanthropic bird species in human, poultry or livestock environments can increase the spread of pathogens and antibiotic-resistant bacteria between wild and domestic animals. We present the first telemetry-based spatial networks for a small songbird. We quantified landscape connectivity exerted by spotless starling movements, and aimed to determine if connectivity patterns were related to carriage of potential pathogens. We captured 28 starlings on a partridge farm in 2020 and tested them for Avian influenza virus, West Nile virus WNV, Avian orthoavulavirus 1, Coronavirus, Salmonella spp. and Escherichia coli. We did not detect any viruses or Salmonella, but one individual had antibodies against WNV or cross-reacting Flaviviruses. We found E. coli in 61 % (17 of 28) of starlings, 76 % (13 of 17) of which were resistant to gentamicin, 12 % (2 of 17) to cefotaxime/enrofloxacin and 6 % (1 of 17) were phenotypic extended spectrum beta-lactamase (ESBL) carriers. We GPS-tracked 17 starlings and constructed spatial networks showing how their movements (i.e. links) connect different farms with nearby urban and natural habitats (i.e. nodes with different attributes). Using E. coli carriage as a proxy for acquisition/dispersal of bacteria, we found differences across spatial networks constructed for E. coli positive (n = 7) and E. coli negative (n = 9) starlings. We used Exponential Random Graph Models to reveal significant differences between networks. In particular, an urban roost was more connected to other sites by movements of E. coli positive than by movements of E. coli negative starlings. Furthermore, an open pine forest used mainly for roosting was more connected to other sites by movements of E. coli negative than by movements of E. coli positive starlings. Using E. coli as a proxy for a potential pathogen carried by starlings, we reveal the pathways of spread that starlings could provide between farms, urban and natural habitats.

Use of dairies by postreproductive flocks of European starlings.

Knowledge of the behavior and movement patterns of European starlings (Sturnus vulgaris L.) is important to wildlife managers that seek to resolve conflicts at livestock facilities. We captured and radio tagged 10 starlings at each of 5 dairies in northeastern Ohio. From September 19 to October 31, 2007, we obtained sufficient data from 40 birds to study their behavior and movements. The birds visited the dairies where they were initially captured (home sites) on 85% of the days, spending 58% of each day at the dairies. Onsite arrival and departure times were 2.5h after sunrise and 3.1h before sunset. Daily visits by radio-tagged cohorts from the other dairies were greatest for the 2 most proximate dairies (1.3 km apart), with number of visits between this pairing >7× that of the 9 other pairings combined (4.1-6.5 km apart). Two birds used their home sites intermittently as roosts, arriving 3.8h before sunset and departing 0.2h after sunrise. In addition to using home-site roosts, these birds also used a distant roost (22km) that was used by 36 of the 40 birds. The efficacy of starling management programs, especially lethal management, depends on degree of site fidelity, use of other facilities, and roosting behavior. For example, starlings that use dairies as roosting sites may require a different management strategy than required at dairies used as daytime sites because of differences in arrival and departure behavior. Our research will help resource managers evaluate current management strategies already in place and change them, if needed, to fit the behavior profile of starlings using dairies and other types of livestock facilities.

Multiple-locus variable-nucleotide tandem repeat subtype analysis implicates European starlings as biological vectors for Escherichia coli O157:H7 in Ohio, USA.

AIMS: To provide molecular epidemiological evidence of avian transmission of Escherichia coli O157:H7 between dairy farms in Ohio, this study was designed to identify genetic relatedness between isolates originating from bovine faecal samples and intestinal contents of European starlings captured on these farms. METHODS AND RESULTS: During a three-year period (2007-2009), cattle (n = 9000) and starlings (n = 430) on 150 different dairy farms in northern Ohio were sampled for the presence of E. coli O157:H7. Isolates were subjected to multiple-locus variable-nucleotide tandem repeat analysis (MLVA). Distinct allelic groups were identified on most farms; however, isolates clustering into three MLVA groups originated from both cattle and birds on different farms. CONCLUSIONS: Sharing of indistinguishable epidemiologically linked E. coli O157 MLVA subtypes between starlings and cattle on different farms supports the hypothesis that these birds contribute to the transmission of E. coli O157:H7 between dairy farms. SIGNIFICANCE AND IMPACT OF STUDY: A continued need exists to identify and to improve preharvest measures for controlling E. coli O157:H7. Controlling wildlife intrusion, particularly European starlings, on livestock operations, may be an important strategy for reducing dissemination of E. coli O157:H7 between farms and thereby potentially decreasing the on-farm prevalence of E. coli O157:H7 and enhancing the safety of the food supply.

Bird-livestock interactions associated with increased cattle fecal shedding of ciprofloxacin-resistant Escherichia coli within feedlots in the United States.

This research study was conducted to determine if bird depredation in feedlots is associated with the prevalence of ciprofloxacin-resistant Escherichia coli in cattle and to determine if removal of invasive bird species could be an effective management strategy to help reduce ciprofloxacin-resistant E. coli in cattle within the United States. European starlings (Sturnus vulgaris) were collected from feedlots within multiple geographic regions within the United States and European starlings within all regions tested positive for ciprofloxacin-resistant E. coli, but prevalence differed by region. Total number of birds on feedlots were positively associated with increased cattle fecal shedding of ciprofloxacin-resistant E. coli. Targeted control of invasive European starlings reduced bird numbers on feedlots by 70.4%, but decreasing populations of European starlings was not associated with corresponding reductions in bovine fecal prevalence of ciprofloxacin-resistant E. coli. These data provide evidence for the role of wild bird depredation in feedlots contributing to fecal shedding of ciprofloxacin-resistant E. coli, but a single month of European starling control in feedlots was not sufficient to impact the fecal carriage of this organism in cattle.

The Role of European Starlings (Sturnus vulgaris) in the Dissemination of Multidrug-Resistant Escherichia coli among Concentrated Animal Feeding Operations.

Antimicrobial use in livestock production is a driver for the development and proliferation of antimicrobial resistance (AMR). Wildlife interactions with livestock, acquiring associated AMR bacteria and genes, and wildlife's subsequent dispersal across the landscape are hypothesized to play an important role in the ecology of AMR. Here, we examined priority AMR phenotypes and genotypes of Escherichia coli isolated from the gastrointestinal tracts of European starlings (Sturnus vulgaris) found on concentrated animal feeding operations (CAFOs). European starlings may be present in high numbers on CAFOs (>100,000 birds), interact with urban environments, and can migrate distances exceeding 1,500 km in North America. In this study, 1,477 European starlings from 31 feedlots in five U.S. states were sampled for E. coli resistant to third generation cephalosporins (3G-C) and fluoroquinolones. The prevalence of 3G-C and fluoroquinolone-resistant E. coli was 4% and 10%, respectively. Multidrug resistance in the E. coli isolates collected (n = 236) was common, with the majority of isolates displaying resistance to six or more classes of antibiotics. Genetic analyses of a subset of these isolates identified 94 genes putatively contributing to AMR, including seven class A and C ß-lactamases as well as mutations in gyrA and parC recognized to confer resistance to quinolones. Phylogenetic and subtyping assessments showed that highly similar isolates (≥99.4% shared core genome, ≥99.6% shared coding sequence) with priority AMR were found in birds on feedlots separated by distances exceeding 150 km, suggesting that European starlings could be involved in the interstate dissemination of priority AMR bacteria.

Dynamics of Campylobacter colonization of a natural host, Sturnus vulgaris (European starling).

Wild European Starlings (Sturnus vulgaris) shed Campylobacter at high rates, suggesting that they may be a source of human and farm animal infection. A survey of Campylobacter shedding of 957 wild starlings was undertaken by culture of faecal specimens and genetic analysis of the campylobacters isolated: shedding rates were 30.6% for Campylobacter jejuni, 0.6% for C. coli and 6.3% for C. lari. Genotyping by multilocus sequence typing (MLST) and antigen sequence typing established that these bacteria were distinct from poultry or human disease isolates with the ST-177 and ST-682 clonal complexes possibly representing starling-adapted genotypes. There was seasonal variation in both shedding rate and genotypic diversity, both exhibiting a maximum during the late spring/early summer. Host age also affected Campylobacter shedding, which was higher in younger birds, and turnover was rapid with no evidence of cross-immunity among Campylobacter species or genotypes. In nestlings, C. jejuni shedding was evident from 9 days of age but siblings were not readily co-infected. The dynamics of Campylobacter infection of starlings differed from that observed in commercial poultry and consequently there was no evidence that wild starlings represent a major source of Campylobacter infections of food animals or humans.

Comparison of Campylobacter populations in wild geese with those in starlings and free-range poultry on the same farm.

Wild geese are a potential source of Campylobacter infection for humans and farm animals and have been implicated in at least two large waterborne disease outbreaks. There have been few investigations into the population biology of Campylobacter in geese, carriage rates are reported to vary (0 to 100%), and no genetic characterization of isolates has been performed. Fecal samples collected from wild geese in Oxfordshire, United Kingdom, were culture positive for C. jejuni (50.2%) and C. coli (0.3%). The C. jejuni (n = 166) isolates were characterized by using multilocus sequence typing and were compared with isolates collected from free-range broiler chickens and wild starlings sampled at the same location. A total of 38 STs, six clonal complexes, and 23 flaA SVR nucleotide STs were identified. The ST-21 and ST-45 complexes (5.4% of isolates) were the only complexes to be identified among isolates from the geese and the other bird species sampled in the same location. These clonal complexes were also identified among human disease isolates collected in the same health care region. The results indicate that large numbers of wild geese carry Campylobacter; however, there was limited mixing of Campylobacter populations among the different sources examined, and the host source could be predicted with high probability from the allelic profile of a C. jejuni isolate. In conclusion, genotypes of C. jejuni isolated from geese are highly host specific, and a comparison with isolates from Oxfordshire cases of human disease revealed that while geese cannot be excluded as a source of infection for humans and farm animals, their contribution is likely to be minor.

Environmental Factors and Zoonotic Pathogen Ecology in Urban Exploiter Species.

Knowledge of pathogen ecology, including the impacts of environmental factors on pathogen and host dynamics, is essential for determining the risk that zoonotic pathogens pose to people. This review synthesizes the scientific literature on environmental factors that influence the ecology and epidemiology of zoonotic microparasites (bacteria, viruses and protozoa) in globally invasive urban exploiter wildlife species (i.e., rock doves [Columba livia domestica], European starlings [Sturnus vulgaris], house sparrows [Passer domesticus], Norway rats [Rattus norvegicus], black rats [R. rattus] and house mice [Mus musculus]). Pathogen ecology, including prevalence and pathogen characteristics, is influenced by geographical location, habitat, season and weather. The prevalence of zoonotic pathogens in mice and rats varies markedly over short geographical distances, but tends to be highest in ports, disadvantaged (e.g., low income) and residential areas. Future research should use epidemiological approaches, including random sampling and robust statistical analyses, to evaluate a range of biotic and abiotic environmental factors at spatial scales suitable for host home range sizes. Moving beyond descriptive studies to uncover the causal factors contributing to uneven pathogen distribution among wildlife hosts in urban environments may lead to targeted surveillance and intervention strategies. Application of this knowledge to urban maintenance and planning may reduce the potential impacts of urban wildlife-associated zoonotic diseases on people.

Nest Material Shapes Eggs Bacterial Environment.

Selective pressures imposed by pathogenic microorganisms to embryos have selected in hosts for a battery of antimicrobial lines of defenses that includes physical and chemical barriers. Due to the antimicrobial properties of volatile compounds of green plants and of chemicals of feather degrading bacteria, the use of aromatic plants and feathers for nest building has been suggested as one of these barriers. However, experimental evidence suggesting such effects is scarce in the literature. During two consecutive years, we explored experimentally the effects of these nest materials on loads of different groups of bacteria (mesophilic bacteria, Enterobacteriaceae, Staphylococcus and Enterococcus) of eggshells in nests of spotless starlings (Sturnus unicolor) at the beginning and at the end of the incubation period. This was also explored in artificial nests without incubation activity. We also experimentally increased bacterial density of eggs in natural and artificial nests and explored the effects of nest lining treatments on eggshell bacterial load. Support for the hypothetical antimicrobial function of nest materials was mainly detected for the year and location with larger average values of eggshell bacterial density. The beneficial effects of feathers and plants were more easily detected in artificial nests with no incubation activity, suggesting an active role of incubation against bacterial colonization of eggshells. Pigmented and unpigmented feathers reduced eggshell bacterial load in starling nests and artificial nest boxes. Results from artificial nests allowed us to discuss and discard alternative scenarios explaining the detected association, particularly those related to the possible sexual role of feathers and aromatic plants in starling nests. All these results considered together confirm the antimicrobial functionality mainly of feathers but also of plants used as nest materials, and highlight the importance of temporally and geographically environmental variation associated with risk of bacterial proliferation determining the strength of such effects. Because of costs associated to nest building, birds should adjust nest building effort to expected bacterial environments during incubation, a prediction that should be further explored.

Mechanisms of antimicrobial resistant Salmonella enterica transmission associated with starling-livestock interactions.

Bird-livestock interactions have been implicated as potential sources for bacteria within concentrated animal feeding operations (CAFO). European starlings (Sturnus vulgaris) in particular are known to contaminate cattle feed and water with Salmonella enterica through their fecal waste. We propose that fecal waste is not the only mechanisms through which starlings introduce S. enterica to CAFO. The goal of this study was to assess if starlings can mechanically move S. enterica. We define mechanical movement as the transportation of media containing S. enterica, on the exterior of starlings within CAFO. We collected 100 starlings and obtained external wash and gastrointestinal tract (GI) samples. We also collected 100 samples from animal pens. Within each pen we collected one cattle fecal, feed, and water trough sample. Isolates from all S. enterica positive samples were subjected to antimicrobial susceptibility testing. All sample types, including 17% of external starling wash samples, contained S. enterica. All sample types had at least one antimicrobial resistant (AMR) isolate and starling GI samples harbored multidrug resistant S. enterica. The serotypes isolated from the starling external wash samples were all found in the farm environment and 11.8% (2/17) of isolates from positive starling external wash samples were resistant to at least one class of antibiotics. This study provides evidence of a potential mechanism of wildlife introduced microbial contamination in CAFO. Mechanical movement of microbiological hazards, by starlings, should be considered a potential source of bacteria that is of concern to veterinary, environmental and public health.

Molecular characterization of Salmonella enterica isolates associated with starling-livestock interactions.

Bird-livestock interactions have been implicated as potential sources for bacteria within concentrated animal feeding operations (CAFO). In this study we characterized XbaI-digested genomic DNA from Salmonella enterica using pulsed-field gel electrophoresis (PFGE). The PFGE analysis was conducted using 182 S. enterica isolates collected from a single CAFO between 2009 and 2012. Samples collected in 2012 were subjected to antimicrobial susceptibility testing. The analysis was limited to S. enterica serotypes, with at least 10 isolates, known to occur in both European starlings (Sturnus vulgaris) and cattle (Bos taurus) within this CAFO. A total of five different serotypes were screened; S. Anatum, S. Kentucky, S. Meleagridis, S. Montevideo, S. Muenchen. These samples were recovered from five different sample types; starling gastrointestinal tracts (GI), starling external wash, cattle feces, cattle feed and cattle water troughs. Indistinguishable S. enterica PFGE profiles were recovered from isolates originating in all sample types. Antimicrobial resistance (AMR) was also associated with indistinguishable S. enterica isolates recovered from all samples types. These data suggests that AMR S. enterica is transmitted between cattle and starlings and that shared feed sources are likely contributing to infections within both species. Moreover we isolated indistinguishable PFGE profiles across all years of data collection, suggesting long-term environmental persistence may be mediated by starling visits to CAFO.

Spatial epidemiology of Escherichia coli O157:H7 in dairy cattle in relation to night roosts Of Sturnus vulgaris (European Starling) in Ohio, USA (2007-2009).

The goal of our study was to use spatial scan statics to determine whether the night roosts of European starlings (Sturnus vulgaris) act as point sources for the dissemination of Escherichia coli O157:H7 among dairy farms. From 2007 to 2009, we collected bovine faecal samples (n = 9000) and starling gastrointestinal contents (n = 430) from 150 dairy farms in northeastern Ohio, USA. Isolates of E. coli O157:H7 recovered from these samples were subtyped using multilocus variable-number tandem repeat analysis (MLVA). Generated MLVA types were used to construct a dendrogram based on a categorical multistate coefficient and unweighted pair-group method with arithmetic mean (UPGMA). Using a focused spatial scan statistic, we identified statistically significant spatial clusters among dairy farms surrounding starling night roosts, with an increased prevalence of E. coli O157:H7-positive bovine faecal pats, increased diversity of distinguishable MLVA types and a greater number of isolates with MLVA types from bovine-starling clades versus bovine-only clades. Thus, our findings are compatible with the hypothesis that starlings have a role in the dissemination of E. coli O157:H7 among dairy farms, and further research into starling management is warranted.

Campylobacter jejuni colonization and population structure in urban populations of ducks and starlings in New Zealand.

A repeated cross-sectional study was conducted to determine the prevalence of Campylobacter spp. and the population structure of C. jejuni in European starlings and ducks cohabiting multiple public access sites in an urban area of New Zealand. The country's geographical isolation and relatively recent history of introduction of wild bird species, including the European starling and mallard duck, create an ideal setting to explore the impact of geographical separation on the population biology of C. jejuni, as well as potential public health implications. A total of 716 starling and 720 duck fecal samples were collected and screened for C. jejuni over a 12 month period. This study combined molecular genotyping, population genetics and epidemiological modeling and revealed: (i) higher Campylobacter spp. isolation in starlings (46%) compared with ducks (30%), but similar isolation of C. jejuni in ducks (23%) and starlings (21%), (ii) significant associations between the isolation of Campylobacter spp. and host species, sampling location and time of year using logistic regression, (iii) evidence of population differentiation, as indicated by FST , and host-genotype association with clonal complexes CC ST-177 and CC ST-682 associated with starlings, and clonal complexes CC ST-1034, CC ST-692, and CC ST-1332 associated with ducks, and (iv) greater genetic diversity and genotype richness in ducks compared with starlings. These findings provide evidence that host-associated genotypes, such as the starling-associated ST-177 and ST-682, represent lineages that were introduced with the host species in the 19th century. The isolation of sequence types associated with human disease in New Zealand indicate that wild ducks and starlings need to be considered as a potential public health risk, particularly in urban areas.

Association of wild bird density and farm management factors with the prevalence of E. coli O157 in dairy herds in Ohio (2007-2009).

Our objective was to determine the role that European starlings (Sturnus vulgaris) play in the epidemiology of Escherichia coli O157:H7 in dairy cattle. We visited 150 dairy farms in Ohio twice during summer and fall months from 2007 to 2009. Fresh faecal pats from 30 lactating cows were collected during each visit. Information on farm management and environmental variables was gathered through a questionnaire administered to the farm owner. The number of starlings observed on the farm was also recorded. Approximately 1% of dairy cattle and 24% of farms were positive for E. coli O157. Risk factors associated with the presence of E. coli O157 in faecal pats included contact between adult cattle and calves, types or number of ventilation and manure management systems and number of birds per milking cow.

Brood size modifications affect plumage bacterial assemblages of European starlings.

During reproduction, birds face trade-offs between time and energy devoted to parental effort and traits associated with self-maintenance. We manipulated brood sizes to investigate the effects of such trade-offs on feather bacterial densities and the structure of bacterial assemblages on feathers in adult European starlings, Sturnus vulgaris, and in vitro feather degradation. As predicted by a trade-off between parental effort and self-maintenance, we found that birds with enlarged broods had more free-living bacteria on their feathers than birds with reduced broods. Furthermore, we found a significant interaction between brood manipulation and original brood size on free-living bacterial densities suggesting that the trade-off is mediated by the adults' initial reproductive investment. In contrast, brood size manipulations had no significant effect on densities of attached bacteria. Using ribosomal intergenic spacer analysis (RISA), we demonstrated that brood manipulations significantly modified the structure (band pattern) of feather-degrading bacterial assemblages, but had no significant effect on their richness (number of bands) or the in vitro feather degradation. In vitro feather degradation varied in relation to the premanipulation brood size and positively with the richness of the feather degrading bacterial community. Besides brood manipulation effect, we found that ecological factors and individual traits, such as the age, the nest location or the capture date, shaped bacterial assemblages and feather degradation capacities.

Verocytotoxin-producing Escherichia coli in wild birds and rodents in close proximity to farms.

Wild animals living close to cattle and pig farms (four each) were examined for verocytotoxin-producing Escherichia coli (VTEC; also known as Shiga toxin-producing E. coli). The prevalence of VTEC among the 260 samples from wild animals was generally low. However, VTEC isolates from a starling (Sturnus vulgaris) and a Norway rat (Rattus norvegicus) were identical to cattle isolates from the corresponding farms with respect to serotype, virulence profile, and pulsed-field gel electrophoresis type. This study shows that wild birds and rodents may become infected from farm animals or vice versa, suggesting a possible role in VTEC transmission.