RESUMO
Shiga toxin-producing Escherichia coli (STEC) and Campylobacter jejuni are notable health hazards associated with the consumption of raw milk. These bacteria may colonize the intestines of asymptomatic cattle and enter bulk tank milk via fecal contamination during milking. We studied the frequency of STEC O157:H7 and C. jejuni contamination in tank milk (n = 785) and the in-line milk filters of milking machines (n = 631) versus the frequency of isolation from cattle feces (n = 257) on three Finnish dairy farms for 1 year. Despite simultaneous isolation of STEC O157:H7 (17%) or C. jejuni (53%) from cattle, these bacteria were rarely isolated from milk filters (2% or <1%, respectively) and milk (0%). As revealed by phylogenomics, one STEC O157:H7 strain at a time was detected on each farm and persisted for ≤12 months despite rigorous hygienic measures. C. jejuni strains of a generalist sequence type (ST-883 and ST-1080) persisted in the herds for ≥11 months, and several other C. jejuni types were detected sporadically. The stx gene carried by STEC was detected more frequently from milk filters (37%) than from milk (7%), suggesting that milk filters are more suitable sampling targets for monitoring than milk. A questionnaire of on-farm practices suggested lower stx contamination of milk when major cleansing in the barn, culling, or pasturing of dairy cows was applied, while a higher average outdoor temperature was associated with higher stx contamination. Because pathogen contamination occurred despite good hygiene and because pathogen detection from milk and milk filters proved challenging, we recommend heat treatment for raw milk before consumption.IMPORTANCE The increased popularity of raw milk consumption has created demand for relaxing legislation, despite the risk of contamination by pathogenic bacteria, notably STEC and C. jejuni However, the epidemiology of these milk-borne pathogens on the herd level is still poorly understood, and data are lacking on the frequency of milk contamination on farms with cattle shedding these bacteria in their feces. This study suggests (i) that STEC contamination in milk can be reduced, but not prevented, by on-farm hygienic measures while fecal shedding is observable, (ii) that milk filters are more suitable sampling targets for monitoring than milk although pathogen detection from both sample matrices may be challenging, and (iii) that STEC and C. jejuni genotypes may persist in cattle herds for several months. The results can be utilized in developing and targeting pathogen monitoring and risk management on the farm level and contributed to the revision of Finnish legislation in 2017.
Assuntos
Campylobacter jejuni/isolamento & purificação , Fezes/microbiologia , Microbiologia de Alimentos , Leite/microbiologia , Escherichia coli Shiga Toxigênica/isolamento & purificação , Animais , Campylobacter jejuni/classificação , Campylobacter jejuni/genética , Bovinos , Indústria de Laticínios/instrumentação , Indústria de Laticínios/métodos , Escherichia coli O157/genética , Escherichia coli O157/isolamento & purificação , Fazendas , Feminino , Finlândia , Genômica , Genótipo , Estudos Longitudinais , Tipagem de Sequências Multilocus , Filogenia , Fatores de Risco , Toxina Shiga/genética , Escherichia coli Shiga Toxigênica/classificação , Escherichia coli Shiga Toxigênica/genética , Sequenciamento Completo do GenomaRESUMO
In August 2016, an outbreak of Shiga toxin-producing Escherichia coli (STEC) and enteropathogenic E. coli (EPEC) with 237 cases occurred in the Helsinki metropolitan area, Finland. Gastroenteritis cases were reported at 11 events served by one catering company. Microbiological and epidemiological investigations suggested rocket salad as the cause of the outbreak. STEC ONT: H11 and EPEC O111:H8 strains isolated from food samples containing rocket were identical to the patient isolates. In this outbreak, the reported symptoms were milder than considered before for STEC infection, and the guidelines for STEC control measures need to be updated based on the severity of the illness. Based on our experience in this outbreak, national surveillance criteria for STEC have been updated to meet the practice in reporting laboratories covering both PCR-positive and culture-confirmed findings. We suggest that EPEC could be added to the national surveillance since diagnostics for EPEC are routinely done in clinical laboratories.
Assuntos
Surtos de Doenças/estatística & dados numéricos , Escherichia coli Enteropatogênica/isolamento & purificação , Infecções por Escherichia coli/epidemiologia , Escherichia coli Shiga Toxigênica/isolamento & purificação , Verduras/microbiologia , Adesinas Bacterianas/genética , Adesinas Bacterianas/metabolismo , Escherichia coli Enteropatogênica/classificação , Escherichia coli Enteropatogênica/genética , Infecções por Escherichia coli/microbiologia , Proteínas de Escherichia coli/genética , Fezes/microbiologia , Finlândia/epidemiologia , Doenças Transmitidas por Alimentos/epidemiologia , Humanos , Reação em Cadeia da Polimerase/métodos , Toxina Shiga/metabolismo , Escherichia coli Shiga Toxigênica/classificação , Escherichia coli Shiga Toxigênica/genéticaRESUMO
In March 2014, a Yersinia pseudotuberculosis (YP) outbreak was detected by a municipal authority in southern Finland. We conducted epidemiological, microbiological and traceback investigations to identify the source. We defined a case as a person with YP infection notified to the National Infectious Disease Registry between February and April 2014, or their household member, with abdominal pain and fever≥38 °C or erythema nodosum. Healthy household members were used as household-matched controls. We identified 43 cases and 50 controls. The illness was strongly associated with the consumption of raw milk from a single producer. The odds ratio of illness increased with the amount of raw milk consumed. Also previously healthy adults became infected by consuming raw milk. Identical YP strains were identified from cases' stool samples, raw milk sampled from a case's refrigerator and from the milk filter at the producer's farm. The producer fulfilled the legal requirements for raw milk production and voluntarily recalled the raw milk and stopped its production. We advised consumers to heat the raw milk to 72 °C for 15 s. Current legislation for raw milk producers should be reviewed and public awareness of health risks linked to raw milk consumption should be increased.
Assuntos
Surtos de Doenças , Leite/microbiologia , Infecções por Yersinia pseudotuberculosis/epidemiologia , Yersinia pseudotuberculosis/isolamento & purificação , Adulto , Animais , Estudos de Casos e Controles , Eletroforese em Gel de Campo Pulsado , Feminino , Finlândia/epidemiologia , Manipulação de Alimentos , Microbiologia de Alimentos , Humanos , Masculino , Razão de Chances , Sorotipagem/métodos , Sequências de Repetição em Tandem , Infecções por Yersinia pseudotuberculosis/diagnóstico , Infecções por Yersinia pseudotuberculosis/transmissãoRESUMO
To prevent foodborne infections from pigs and cattle, the whole food chain must act to minimize the contamination of products, including biosecurity measures which prevent infections via feed and the environment in production farms. Rodents and other small mammals can be reservoirs of and key vectors for transmitting zoonotic bacteria and viruses to farm animals, through direct contact but more often through environmental contamination. In line with One Health concept, we integrated results from a sampling study of small mammals in farm environments and data from a capture-recapture experiment into a probabilistic model which quantifies the degree of environmental exposure of zoonotic bacteria by small mammals to farm premises. We investigated more than 1200 small mammals trapped in and around 38 swine and cattle farm premises in Finland in 2017/2018. Regardless of the farm type, the most common species caught were the yellow-necked mouse (Apodemus flavicollis), bank vole (Clethrionomys glareolus), and house mouse (Mus musculus). Of 554 intestine samples (each pooled from 1 to 10 individuals), 33% were positive for Campylobacter jejuni. Yersinia enterocolitica was detected in 8% of the pooled samples, on 21/38 farm premises. Findings of Salmonella and the Shiga-toxin producing Escherichia coli (STEC) were rare: the pathogens were detected in only single samples from four and six farm premises, respectively. The prevalence of Campylobacter, Salmonella, Yersinia and STEC in small mammal populations was estimated as 26%/13%, 1%/0%, 2%/3%, 1%/1%, respectively, in 2017/2018. The exposure probability within the experimental period of four weeks on farms was 17-60% for Campylobacter and 0-3% for Salmonella. The quantitative model is readily applicable to similar integrative studies. Our results indicate that small mammals increase the risk of exposure to zoonotic bacteria in animal production farms, thus increasing risks also for livestock and human health.
Assuntos
Doenças dos Bovinos , Doenças dos Suínos , Animais , Bovinos , Suínos , Prevalência , Doenças dos Suínos/epidemiologia , Doenças dos Suínos/microbiologia , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/transmissão , Finlândia/epidemiologia , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/microbiologia , Doenças dos Bovinos/transmissão , Roedores/microbiologia , Zoonoses Bacterianas/epidemiologia , Zoonoses Bacterianas/microbiologia , Zoonoses/epidemiologia , Reservatórios de Doenças/veterinária , Reservatórios de Doenças/microbiologia , Medição de Risco , FazendasRESUMO
One chromosomal virulence marker of Yersinia is the gene ail, which encodes Ail, an outer membrane protein that promotes attachment and invasion. A high correlation has been found between the ail gene and the virulence of Yersinia. Here, we report two Yersinia enterocolitica biotype 1A strains that are usually nonpathogenic and carry the ail gene. The ail gene sequences of biotype 1A strains displayed similarity to the bioserotype 1B/O:8 strain 8081. The finding suggests that ail-based detection methods for Y. enterocolitica alone are insufficient to detect real pathogenic strains.
Assuntos
Proteínas da Membrana Bacteriana Externa/genética , DNA Bacteriano/genética , Yersiniose/microbiologia , Yersinia enterocolitica/genética , Sequência de Bases , DNA Ribossômico/química , DNA Ribossômico/genética , Feminino , Finlândia , Humanos , Pessoa de Meia-Idade , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Polimorfismo Genético , RNA Ribossômico 16S/genética , Alinhamento de Sequência , Análise de Sequência de DNA , Virulência/genética , Yersinia enterocolitica/classificação , Yersinia enterocolitica/isolamento & purificação , Yersinia enterocolitica/patogenicidadeRESUMO
Packaged raw milk contaminated with Yersinia pseudotuberculosis mediated a large yersiniosis outbreak in southern Finland in 2014. The outbreak was traced back to a single dairy farm in southern Finland. Here we explore risk factors leading to the outbreak through epidemiologic investigation of the outbreak farm and through genomic and phenotypic characterization of the farm's outbreak and non-outbreak associated Y. pseudotuberculosis strains. We show that the outbreak strain persisted on the farm throughout the 7-month study, whereas the non-outbreak strains occurred sporadically. Phylogenomic analysis illustrated that the outbreak strain was related to previously published genomes of wild animal isolates from Finland, implying that wild animals were a potential source of the outbreak strain to the farm. We observed allelic differences between the farm's outbreak and non-outbreak strains in several genes associated with virulence, stress response and biofilm formation, and found that the outbreak strain formed biofilm in vitro and maintained better growth fitness during cold stress than the non-outbreak strains. Finally, we demonstrate the rapid growth of the outbreak strain in packaged raw milk during refrigerated storage. This study provides insight of the risk factors leading to the Y. pseudotuberculosis outbreak, highlights the importance of pest control to avoid the spread of pathogens from wild to domestic animals, and demonstrates that the cold chain is insufficient as the sole risk management strategy to control Y. pseudotuberculosis risk associated with raw drinking milk.
RESUMO
EN ISO 10273 method for the detection of pathogenic Yersinia enterocolitica in foods was validated in the project Mandate M/381 funded by European Commission. A total of 14 laboratories from five European countries participated in the interlaboratory study (ILS) organized during 2013 and 2014. Before the ILS, the method was revised by an international group of experts and the performance of the revised method was assessed in an ILS study. The results are published as a part of the standard EN ISO 10273 revision. The study included three rounds with different sample types; raw milk, iceberg lettuce and minced meat, inoculated with a low and high level of pathogenic Y. enterocolitica strains representing major pathogenic bioserotypes 4/O:3 and 2/O:9. The homogeneity and stability of the samples were verified before dispatching them to the laboratories. The results demonstrated the method sensitivity of 96% in raw milk, 97% in minced meat, and 98% in lettuce at high inoculation level of pathogenic Y. enterocolitica. The specificity was 100% in raw milk, 96% in minced meat, and 98% in lettuce. The level of detection, LOD50, varied between study rounds, being 9.4â¯CFU/25â¯ml in raw milk, 9.9â¯CFU/25â¯g in minced meat and 63â¯CFU/25â¯g in lettuce samples. During the study, confirmation by using real-time PCR method ISO/TS 18867 together with pyrazinamidase testing was also validated, as alternative to conventional biochemical confirmation. When comparing different isolation steps used in the revised method during the study rounds, PSB enrichment and plating on CIN after alkaline (KOH) treatment showed the highest sensitivity (52-92%) in raw milk and minced meat samples. In lettuce samples, however, ITC with KOH treatment before plating on CIN showed higher sensitivity (64% at low level; 82% at high level) than plating on CIN from PSB with KOH treatment (44% at low level; 74% at high level). Statistical analysis of different isolation steps supported the use of two enrichment media, PSB and ITC, in the revised method. Recovery of pathogenic Y. enterocolitica on CIN was most efficient after KOH treatment and, based on the analysis, plating on CIN agar without KOH treatment could be left as optional procedure in the method.
Assuntos
Microbiologia de Alimentos/métodos , Yersinia enterocolitica/fisiologia , Animais , Europa (Continente) , União Europeia , Lactuca/microbiologia , Limite de Detecção , Carne/microbiologia , Leite/microbiologia , Reprodutibilidade dos Testes , Yersinia enterocolitica/isolamento & purificaçãoAssuntos
Surtos de Doenças , Infecções por Yersinia pseudotuberculosis/epidemiologia , Yersinia pseudotuberculosis , Eletroforese em Gel de Campo Pulsado , Finlândia/epidemiologia , Epidemiologia Molecular/métodos , Antígenos O/análise , Sorotipagem , Yersinia pseudotuberculosis/classificação , Yersinia pseudotuberculosis/genéticaRESUMO
Many clinical laboratories are familiar with a sizeable group of "unserotypeable Yersinia enterocolitica" strains. Due to identification problems, this group may hide Y. bercovieri, Y. mollaretii, and Y. rohdei strains. We present a simple scheme to distinguish between pathogenic Y. enterocolitica and potentially nonpathogenic Y. enterocolitica-like strains.
Assuntos
RNA Bacteriano/genética , RNA Ribossômico 16S/genética , Yersinia enterocolitica/classificação , Yersinia enterocolitica/genética , Yersinia/classificação , Yersinia/genética , Técnicas de Tipagem Bacteriana , Finlândia , Genótipo , Humanos , Fenótipo , Reação em Cadeia da Polimerase , Especificidade da Espécie , Virulência , Yersinia/isolamento & purificação , Yersinia/patogenicidade , Yersinia enterocolitica/isolamento & purificação , Yersinia enterocolitica/patogenicidadeRESUMO
BACKGROUND: Outbreaks of Yersinia pseudotuberculosis infection have been epidemiologically linked to fresh produce, but the bacterium has not been recovered from the food items implicated. In May 2003, a cluster of gastrointestinal illness and erythema nodosum was detected among schoolchildren who had eaten lunches prepared by the same institutional kitchen. METHODS: We conducted a case-control study and trace-back, environmental, and laboratory investigations. Case patients had culture-confirmed Y. pseudotuberculosis O:1 infection, erythema nodosum, or reactive arthritis. Bacterial isolates from clinical and environmental samples were compared using pulsed-field gel electrophoresis (PFGE). RESULTS: Of 7392 persons at risk, 111 (1.5%) met the case definition; 76 case patients and 172 healthy control subjects were enrolled in the case-control study. Only raw grated carrots were significantly associated with illness in a logistic-regression model (multivariable odds ratio, 5.7 [95% confidence interval, 1.7-19.5]); a dose response was found for increasing amount of consumption. Y. pseudotuberculosis O:1 isolates from 39 stool specimens and from 5 (42%) of 12 soil samples that contained carrot residue and were obtained from peeling and washing equipment at the production farm were indistinguishable by PFGE. CONCLUSIONS: Carrots contaminated early in the production process caused a large point-source outbreak. Our findings enable the development of evidence-based strategies to prevent outbreaks of this emerging foodborne pathogen.
Assuntos
Daucus carota/microbiologia , Eritema Nodoso/microbiologia , Microbiologia de Alimentos , Gastroenterite/microbiologia , Infecções por Yersinia pseudotuberculosis/epidemiologia , Yersinia pseudotuberculosis/isolamento & purificação , Adolescente , Artrite Reativa/epidemiologia , Artrite Reativa/microbiologia , Estudos de Casos e Controles , Criança , Pré-Escolar , Surtos de Doenças , Eritema Nodoso/epidemiologia , Feminino , Finlândia/epidemiologia , Gastroenterite/epidemiologia , Humanos , Lactente , Recém-Nascido , Masculino , Fatores de TempoRESUMO
Strains (n = 203) of Yersinia species were used in genotyping and PCR experiments in order to evaluate the genotyping potential of the YeO:3RS probe. This probe comprises a 12.5 kb genomic fragment of the Y. enterocolitica O:3 lipopolysaccharide O-antigen gene cluster cloned into plasmid pBR322. The genotyping potential of YeO:3RS was shown to reside in the region upstream of the O-antigen gene cluster, i.e., in the first 1.65 kb of the cloned genomic fragment that contains a repeated sequence (RS) present in multiple copies in the genome. In genotyping, the YeO:3RS probe was hybridised to DNA of Yersinia enterocolitica isolates (n = 112) from humans, animals and food, along with strains of other Yersinia species (n = 5) and Salmonella enterica strains (n = 3). The YeO:3RS probe efficiently detected and subtyped all European pathogenic Yersinia enterocolitica isolates of the serobiotypes O:3/4, O:9/2 and O:5,27/2 studied (n = 87), whereas it hybridised only weakly or not at all with the other strains. Within Yersinia enterocolitica serobiotype O:3/4 strains, YeO:3RS genotyping was as discriminatory as genotyping by pulsed-field gel electrophoresis (PFGE) of XbaI-NotI digested genomic DNA. When these two methods were combined, YeO:3RS genotyping divided both of the two predominant PFGE types into six subtypes, thus increasing the discrimination. In PCR screening of additional 86 Yersinia strains, the 1.65 kb region was detected in European pathogenic serotypes O:1 and O:2 in addition to serotypes O:3, O:5,27 and O:9, indicating that it can be exploited in detecting and typing of European pathogenic serotypes in general.
Assuntos
DNA Bacteriano/genética , Yersiniose/diagnóstico , Yersinia enterocolitica/genética , Southern Blotting , Impressões Digitais de DNA/métodos , Sondas de DNA , DNA Bacteriano/química , Eletroforese em Gel de Campo Pulsado , Humanos , Antígenos O/genética , Plasmídeos , Sequências Repetitivas de Ácido Nucleico , Sorotipagem/métodos , Virulência , Yersiniose/epidemiologia , Yersiniose/microbiologia , Yersinia enterocolitica/classificação , Yersinia enterocolitica/patogenicidadeRESUMO
BACKGROUND: The vehicles and sources of Yersinia pseudotuberculosis infection are unknown. In Finland, clinical microbiology laboratories routinely report Y. pseudotuberculosis isolations and submit isolates for serotype analysis. In October 1998, the number of serotype O:3 infections increased markedly. METHODS: Case patients with culture-confirmed Y. pseudotuberculosis O:3 infection were identified by use of laboratory-based surveillance. We conducted a population-based case-control study. Healthy community control subjects were matched by age, sex, and postal code. Isolates were subtyped by pulsed-field gel electrophoresis (PFGE). RESULTS: Nationwide, 47 case patients were identified (age range, 2-77 years; median, 19 years). One patient with bacteremia died; 5 underwent appendectomies. We enrolled 38 case patients and 76 control subjects in the case-control study. Seventy-one percent of case patients and 42% of control subjects reported having eaten iceberg lettuce (matched odds ratio, 3.8; 95% confidence interval, 1.3-9.4); a dose-response relationship was found for increasing frequency of consumption. Of the 27 isolates obtained from case patients and tested in the analysis, all had indistinguishable PFGE patterns. Four lunch cafeterias that had served iceberg lettuce were associated with clusters of case patients. The lettuce was traced back to originating farms. CONCLUSIONS: Iceberg lettuce was implicated as the vehicle of a widespread foodborne Y. pseudotuberculosis outbreak. Ongoing laboratory-based surveillance and serotype analysis were essential in the rapid detection of infection. Cases of yersiniosis, which appear to be sporadic, may be part of unrecognized outbreaks caused by contaminated fresh produce.