Campylobacter jejuni has caused several campylobacteriosis outbreaks via raw milk consumption. This study reports follow-up of a milk-borne campylobacteriosis outbreak that revealed persistent C. ...jejuni contamination of bulk tank milk for seven months or longer. Only the outbreak-causing strain, representing sequence type (ST) 883, was isolated from milk, although other C. jejuni STs were also isolated from the farm. We hypothesized that the outbreak strain harbors features that aid its environmental transmission or survival in milk. To identify such phenotypic features, the outbreak strain was characterized for survival in refrigerated raw milk and in aerobic broth culture by plate counting and for biofilm formation on microplates by crystal violet staining and quantification. Furthermore, whole-genome sequences were studied for such genotypic features. For comparison, we characterized isolates representing other STs from the same farm and an ST-883 isolate that persisted on another dairy farm, but was not isolated from bulk tank milk. With high inocula (105 CFU/ml), ST-883 strains survived in refrigerated raw milk longer (4-6 days) than the other strains (≤3 days), but the outbreak strain showed no outperformance among ST-883 strains. This suggests that ST-883 strains may share features that aid their survival in milk, but other mechanisms are required for persistence in milk. No correlation was observed between survival in refrigerated milk and aerotolerance. The outbreak strain formed a biofilm, offering a potential explanation for persistence in milk. Whether biofilm formation was affected by pTet-like genomic element and phase-variable genes encoding capsular methyltransferase and cytochrome C551 peroxidase warrants further study. This study suggests a phenotypic target candidate for interventions and genetic markers for the phenotype, which should be investigated further with the final aim of developing control strategies against C. jejuni infections.
Campylobacteriosis is one of the most common causes of bacterial gastroenteritis. However, the clinical course of the illness varies in symptoms and severity. The aim of this study was to ...characterize Campylobacter jejuni (34 isolates) and C. coli (9 isolates) from persons with diarrheal and non-diarrheal stools at the time of examination and fecal sampling, in Poland by using whole-genome sequencing (WGS). Multilocus sequence typing (MLST) analysis revealed a high diversity with a total of 20 sequence types (STs) among 26 Campylobacter isolates from diarrheic and 13 STs among 17 isolates from non-diarrheic persons. ST-50 and ST-257 were most common in both groups. The phenotypic resistance rate was 74.4% for ciprofloxacin, 67.4% for sulfamethoxazole/trimethoprim, 58.1% for amoxicillin, 48.8% for tetracycline, and 46.5% for ceftriaxone. Only single isolates were resistant to erythromycin, gentamicin, and amoxicillin/clavulanic acid. Overall genotypic resistance toward amoxicillin, fluoroquinolones, tetracyclines, and aminoglycosides was predicted to occur in 93.1, 67.4, 48.8, and 11.6% of the isolates, respectively. None of the isolates showed the presence of the erm(B) gene or mutation in 23S rRNA. Neither was variation found in the important target region in L4 and L22 ribosomal proteins. In regard to the CmeABC efflux pump, a set of variable mutations affecting the regulatory region was noted. All Campylobacter isolates possessed genes associated with adhesion (cadF, jlpA, porA, and pebA) and invasion (ciaB, pldA, and flaC). The type IV secretion system (T4SS) was found in isolates from both diarrheic (15.4%, CI 95%: 6.1-33.5%) and non-diarrheic (23.5%, CI 95%: 9.6-47.3%) persons. The rates of the presence of cytolethal distending toxin cdtABC gene cluster and type VI secretion system (T6SS) were higher in Campylobacter isolates obtained from persons with diarrhea (96.2%, CI 95%: 81.7-99.3% and 26.9%, CI 95%: 13.7-46.1%) compared to isolates from non-diarrheic persons (76.5%, CI 95%: 52.7-90.4% and 11.8%, CI 95%: 3.3-34.3%). The lack of statistically significant differences between two groups in tested virulence factors suggests that individual susceptibility of the host might play more determining role in the disease outcome than characteristics of the infecting strain.Campylobacteriosis is one of the most common causes of bacterial gastroenteritis. However, the clinical course of the illness varies in symptoms and severity. The aim of this study was to characterize Campylobacter jejuni (34 isolates) and C. coli (9 isolates) from persons with diarrheal and non-diarrheal stools at the time of examination and fecal sampling, in Poland by using whole-genome sequencing (WGS). Multilocus sequence typing (MLST) analysis revealed a high diversity with a total of 20 sequence types (STs) among 26 Campylobacter isolates from diarrheic and 13 STs among 17 isolates from non-diarrheic persons. ST-50 and ST-257 were most common in both groups. The phenotypic resistance rate was 74.4% for ciprofloxacin, 67.4% for sulfamethoxazole/trimethoprim, 58.1% for amoxicillin, 48.8% for tetracycline, and 46.5% for ceftriaxone. Only single isolates were resistant to erythromycin, gentamicin, and amoxicillin/clavulanic acid. Overall genotypic resistance toward amoxicillin, fluoroquinolones, tetracyclines, and aminoglycosides was predicted to occur in 93.1, 67.4, 48.8, and 11.6% of the isolates, respectively. None of the isolates showed the presence of the erm(B) gene or mutation in 23S rRNA. Neither was variation found in the important target region in L4 and L22 ribosomal proteins. In regard to the CmeABC efflux pump, a set of variable mutations affecting the regulatory region was noted. All Campylobacter isolates possessed genes associated with adhesion (cadF, jlpA, porA, and pebA) and invasion (ciaB, pldA, and flaC). The type IV secretion system (T4SS) was found in isolates from both diarrheic (15.4%, CI 95%: 6.1-33.5%) and non-diarrheic (23.5%, CI 95%: 9.6-47.3%) persons. The rates of the presence of cytolethal distending toxin cdtABC gene cluster and type VI secretion system (T6SS) were higher in Campylobacter isolates obtained from persons with diarrhea (96.2%, CI 95%: 81.7-99.3% and 26.9%, CI 95%: 13.7-46.1%) compared to isolates from non-diarrheic persons (76.5%, CI 95%: 52.7-90.4% and 11.8%, CI 95%: 3.3-34.3%). The lack of statistically significant differences between two groups in tested virulence factors suggests that individual susceptibility of the host might play more determining role in the disease outcome than characteristics of the infecting strain.
Campylobacter spp., especially C. jejuni, is the most common zoonotic pathogen in humans worldwide. In Nordic countries, the prevalence of C. jejuni in broilers, which is an important reservoir of ...human infections, is generally low. Thus, other sources of domestically acquired infections besides chicken meat need to be considered. Game birds are known to carry a variety of zoonotic agents, including Campylobacter. The aim of this study was to investigate Campylobacter spp. carriage in a flock of reared pheasants at hunting in two successive samplings to better understand the dynamics of Campylobacter infections in pheasants. Overall, 72% of the intestinal samples were positive for Campylobacter spp. by direct culture on mCCDA. C. jejuni was the only species identified. The isolates were genotyped using whole genome sequencing (WGS), multilocus sequence typing (MLST), and ad hoc whole genome MLST (wgMLST). Two distinct C. jejuni clones were identified among the 18 isolates studied, representing MLST sequence types (STs) ST-45 and ST-699. The ST-45 isolates were closely related to previous human clinical isolates using core genome MLST (cgMLST). In contrast, the ST-699 isolates forming the dominant clone in the latter sampling were quite distinct from previously described cgMLST profiles from different hosts and sources worldwide. In conclusion, the intestine of reared pheasants is commonly colonized by C. jejuni and may carry genotypes relevant to infections in livestock and humans. Hygienic measures are needed to limit the spread of infection in reared flocks. Especially farmers and hunters having direct contact with pheasant offal need to be aware of the associated zoonosis risk to protect themselves and their working dogs alike. Biosecurity measures to improve the safety and reduce the zoonosis risk associated with pheasant farming should be further investigated.
Campylobacter jejuni is the most common bacterial cause of human gastroenteritis worldwide. Due to the sporadic nature of infection, sources often remain unknown. Multilocus sequence typing (MLST) ...has been successfully applied to population genetics of Campylobacter jejuni and mathematical modelling can be applied to the sequence data. Here, we analysed the population structure of a total of 250 Finnish C. jejuni isolates from bovines, poultry meat and humans collected in 2003 using a combination of Bayesian clustering (BAPS software) and phylogenetic analysis.
In the first phase we analysed sequence types (STs) of 102 Finnish bovine C. jejuni isolates by MLST and found a high diversity totalling 50 STs of which nearly half were novel. In the second phase we included MLST data from domestic human isolates as well as poultry C. jejuni isolates from the same time period. Between the human and bovine isolates we found an overlap of 72.2%, while 69% of the human isolates were overlapping with the chicken isolates. In the BAPS analysis 44.3% of the human isolates were found in bovine-associated BAPS clusters and 45.4% of the human isolates were found in the poultry-associated BAPS cluster. BAPS reflected the phylogeny of our data very well.
These findings suggest that bovines and poultry were equally important as reservoirs for human C. jejuni infections in Finland in 2003. Our results differ from those obtained in other countries where poultry has been identified as the most important source for human infections. The low prevalence of C. jejuni in poultry flocks in Finland could explain the lower attribution of human infection to poultry. Of the human isolates 10.3% were found in clusters not associated with any host which warrants further investigation, with particular focus on waterborne transmission routes and companion animals.
Abstract Background European hedgehogs ( Erinaceus europaeus) are widely distributed across Europe. They may play an important role by spreading zoonotic bacteria in the environment and to humans and ...animals. The aim of our work was to study the prevalence and characteristics of the most important foodborne bacterial pathogens in wild hedgehogs. Results Faecal samples from 148 hospitalised wild hedgehogs originating from the Helsinki region in southern Finland were studied. Foodborne pathogens were detected in 60% of the hedgehogs by PCR. Listeria (26%) and STEC (26%) were the most common foodborne pathogens. Salmonella, Yersinia , and Campylobacter were detected in 18%, 16%, and 7% of hedgehogs, respectively. Salmonella and Yersinia were highly susceptible to the tested antimicrobials. Salmonella Enteritidis and Listeria monocytogenes 2a were the most common types found in hedgehogs. All S. Enteritidis belonged to one sequence type (ST11), forming four clusters of closely related isolates. L. monocytogenes was genetically more diverse than Salmonella , belonging to 11 STs. C. jejuni ST45 and ST677, Y. pseudotuberculosis O:1 of ST9 and ST42, and Y. enterocolitica O:9 of ST139 were also found. Conclusions Our study shows that wild European hedgehogs should be considered an important source of foodborne pathogens, and appropriate hygiene measures after any contact with hedgehogs and strict biosecurity around farms are therefore important.
Poultry are considered a major reservoir and source of human campylobacteriosis, but the roles of environmental reservoirs, including wild birds, have not been assessed in depth. In this study, we ...isolated and characterized
from western jackdaws (
= 91, 43%), mallard ducks (
= 82, 76%), and pheasants (
= 9, 9%). Most of the western jackdaw and mallard duck
isolates represented multilocus sequence typing (MLST) sequence types (STs) that diverged from those previously isolated from human patients and various animal species, whereas all pheasant isolates represented ST-19, a common ST among human patients and other hosts worldwide. Whole-genome MLST revealed that mallard duck ST-2314 and pheasant ST-19 isolates represented bacterial clones that were genetically highly similar to human isolates detected previously. Further analyses revealed that in addition to a divergent ClonalFrame genealogy, certain genomic characteristics of the western jackdaw
isolates, e.g., a novel
gene cluster and the type VI secretion system (T6SS), may affect their host specificity and virulence. Game birds may thus pose a risk for acquiring campylobacteriosis; therefore, hygienic measures during slaughter and meat handling warrant special attention.
The roles of environmental reservoirs, including wild birds, in the molecular epidemiology of
have not been assessed in depth. Our results showed that game birds may pose a risk for acquiring campylobacteriosis, because they had
genomotypes highly similar to human isolates detected previously. Therefore, hygienic measures during slaughter and meat handling warrant special attention. On the contrary, a unique phylogeny was revealed for the western jackdaw isolates, and certain genomic characteristics identified among these isolates are hypothesized to affect their host specificity and virulence. Comparative genomics within sequence types (STs), using whole-genome multilocus sequence typing (wgMLST), and phylogenomics are efficient methods to analyze the genomic relationships of
isolates.
Wastewater discharge and runoff waters are significant sources of human and animal fecal microbes in surface waters. Human-derived fecal contamination of water is generally estimated to pose a ...greater risk to human health than animal fecal contamination, but animals may serve as reservoirs of zoonotic pathogens. In this study, quantitative microbial risk assessment (QMRA) tools were used to evaluate the hygienic impact of sewage effluents and runoff water from municipalities and animal farms on surface and bathing waters. The human-specific microbial source tracking (MST) marker HF183 was used to evaluate the dilution of fecal pathogens originating from the sewage effluent discharge to the downstream watershed. As novel risk management options, the efficiency of UV-LED disinfection and wetland treatment as well as biochar filtration was tested on-site for the contamination sources. According to the dilution pattern of the MST marker HF183, microbes from wastewater were diluted (2.3-3.7 log10) in the receiving waters. The scenario-based QMRA revealed, that the health risks posed by exposure to human-specific norovirus GII and zoonotic Campylobacter jejuni during the bathing events were evaluated. The risk for gastroenteritis was found to be elevated during wastewater contamination events, where especially norovirus GII infection risk increased (1-15 cases per day among 50 bathers) compared with the business as usual (BAU) situation (1 case per day). The noted C. jejuni infection risk was associated with animal farm contamination (1 case per day, versus 0.2-0.6 cases during BAU). Tertiary treatment of wastewater with wetland treatment and UV-LED disinfection effectively reduced the waterborne gastroenteritis risks associated with bathing. Based on the experiences from this study, a QMRA-based approach for health risk evaluations at bathing sites can be useful and is recommended for bathing site risk assessments in the future. In case of low pathogen numbers at the exposure sites, the MST marker HF183 could be used as a pathogen dilution coefficient for the watershed under evaluation. The full-scale implementation of novel tertiary treatment options at wastewater treatment plants (WWTPs) as well as on-site runoff water treatment options should be considered for infection risk management at locations where scenario-based QMRA implies elevated infection risks.
(
) is the most common cause of human bacterial gastroenteritis in the world. Food-borne campylobacteriosis is thought to be commonly caused by the handling and consumption of undercooked chicken ...meat, but the epidemiology of this disease is complex and remains poorly characterized, especially in the Nordic countries. Here, we used state-of-the-art methods in genetic epidemiology combined with patient background and temporal association data to trace domestically acquired human
infections (
= 50) to chicken meat, in a midsize Nordic town in Finland during a seasonal peak. Although 59.2% of the human isolates shared a sequence type (ST) with a chicken batch slaughtered prior to the onset of disease, further analysis at the whole-genome level (core genome and whole-genome multilocus sequence typing, cgMLST and wgMLST, respectively) traced a mere nine cases (18.4%) to fresh chicken meat. Human isolates also shared genotypes with isolates collected from chicken batches slaughtered after the onset of the human disease, highlighting the role of alternative transmission pathways from chickens to humans besides the food chain, or a shared third source. The high resolution offered by wgMLST, combined with simple metadata, offers a more accurate way to trace sporadic cases to possible sources and reveal disseminated outbreak clustering in time, confirming the importance of complementing epidemiological investigations with molecular epidemiological data.
There has been a significant increase in the number of reported human cryptosporidiosis cases in recent years. The aim of this study is to estimate the prevalence of Cryptosporidium spp. in wild ...rodents and shrews, and investigate the species and genotype distribution to assess zoonotic risk. Partial 18S rRNA gene nested-PCR reveals that 36.8, 53.9 and 41.9% of mice, voles and shrews are infected with Cryptosporidium species. The highest prevalence occurred in the Microtus agrestis (field vole) and Myodes glareolus (bank vole). Interestingly, bank voles caught in fields were significantly more often Cryptosporidium-positive compared to those caught in forests. The proportion of infected animals increases from over-wintered (spring and summer) to juveniles (autumn) suggesting acquired immunity in older animals. Based on Sanger sequencing and phylogenetic analyses, Apodemus flavicollis (yellow-necked mouse) is commonly infected with zoonotic C. ditrichi. Voles carry multiple different Cryptosporidium sp. and genotypes, some of which are novel. C. andersoni, another zoonotic species, is identified in the Craseomys rufocanus (grey-sided vole). Shrews carry novel shrew genotypes. In conclusion, this study indicates that Cryptosporidium protozoan are present in mouse, vole and shrew populations around Finland and the highest zoonotic risk is associated with C. ditrichi in Apodemus flavicollis and C. andersoni in Craseomys rufocanus. C. parvum, the most common zoonotic species in human infections, was not detected.
Small mammals are known to carry
spp.; however, little is known about the genotypes and their role in human infections. We studied intestinal content from small wild mammals collected in their ...natural habitats in Finland in 2010-2017, and in close proximity to 40 pig or cattle farms in 2017. The animals were trapped using traditional Finnish metal snap traps.
spp. were isolated from the intestinal content using direct plating on mCCDA. A total of 19% of the captured wild animals (
= 577) and 41% of the pooled farm samples (
= 227) were positive for
, which was the only
species identified. The highest prevalence occurred in yellow-necked mice (
) and bank voles (
) which carried
spp. in 66.3 and 63.9% of the farm samples and 41.5 and 24.4% of individual animals trapped from natural habitats, respectively. Interestingly, all house mouse (
) and shrew (
spp.) samples were negative for
spp.
isolates (
= 145) were further characterized by whole-genome sequencing. Core genome multilocus sequence typing (cgMLST) clustering showed that mouse and vole strains were separated from the rest of the
population (636 and 671 allelic differences, 94 and 99% of core loci, respectively). Very little or no alleles were shared with
genomes described earlier from livestock or human isolates. FastANI results further indicated that
strains from voles are likely to represent a new previously undescribed species or subspecies of
. Core-genome phylogeny showed that there was no difference between isolates originating from the farm and wild captured animals. Instead, the phylogeny followed the host species-association. There was some evidence (one strain each) of livestock-associated
occurring in a farm-caught
and a brown rat (
), indicating that although small mammals may not be the original reservoir of
colonizing livestock, they may sporadically carry
strains occurring mainly in livestock and be associated with disease in humans.
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