Prevalence and risk factors for extended-spectrum β-lactamase or AmpC-producing Escherichia coli in organic dairy herds in the Netherlands.

Extended-spectrum β-lactamase and AmpC-producing Escherichia coli (ESBL/AmpC) are an emerging problem and are hypothesized to be associated with antimicrobial use (AMU), and more specifically with the use of third- and fourth-generation cephalosporins. Whether ESBL/AmpC also occur in organic dairy herds, which have restricted AMU, is not known. Additionally, it is unknown whether, in addition to restricted AMU, other factors in organic herd management are associated with ESBL/AmpC herd status. The aim of this study was to estimate the prevalence of ESBL/AmpC in organic dairy herds in the Netherlands. Subsequently, the relationships between the ESBL/AmpC herd status and AMU and between ESBL/AmpC herd status and farmers' management were assessed in organic dairy herds. For this study, 90 randomly selected, officially registered organic dairy herds were included. The ESBL/AmpC herd status was determined based on the bacteriological culture result of a slurry sample. The sensitivity of testing slurry samples for ESBL/AmpC herd status is less than 100% for detecting herds with a low ESBL/AmpC prevalence. For that reason, herds that tested positive for ESBL/AmpC in slurry were defined as positive and herds with negative slurry samples were defined as unsuspected. A comprehensive questionnaire on management practices was conducted and records on specified antimicrobials that were provided to these herds by the veterinary service providers were obtained. From the data on antimicrobial supplies by the veterinarian, the animal daily defined dose of antimicrobials per farm per year (DDDAF) was calculated. Descriptive statistics were used to describe the relation between the ESBL/AmpC herd status and DDDAF. Multivariable logistic regression models were used to evaluate management factors associated with the ESBL/AmpC herd status. We found ESBL/AmpC in 12 of the 90 (13%; 95% confidence interval=7-22%) slurry samples from organic dairy herds. The median DDDAF in organic dairy herds was 0.5, which was not significantly different between ESBL/AmpC-positive and unsuspected dairy herds. No association could be found between the use of different types of antimicrobials, such as third- and fourth-generation cephalosporins, and ESBL/AmpC herd status. Factors that were associated with higher odds of being ESBL/AmpC-positive were pig farms located within a 2-km radius of the barn, applying parental treatment for clinical mastitis, and providing milk replacer to the female calves after colostrum intake. The prevalence of ESBL/AmpC in organic dairy herds appeared lower than the prevalence in previous studies conducted in conventional dairy herds. Apparently, ESBL/AmpC are also present in herds with low AMU; this indicates that other factors than AMU are also associated with ESBL/AmpC herd status.

Santman-Berends IMGA ，Gonggrijp MA ，Hage JJ ，Heuvelink AE ，Velthuis A ，Lam TJGM ，van Schaik G ... -  《-》

Prevalence and risk factors for extended-spectrum β-lactamase- and AmpC-producing Escherichia coli in dairy farms.

A cross-sectional study was conducted to evaluate the prevalence of extended-spectrum β-lactamase (ESBL)- and plasmid-mediated AmpC-producing Escherichia coli and associated risk factors in dairy herds. One hundred dairy herds were randomly selected and sampled to study the presence of ESBL- and AmpC-producing E. coli in slurry samples. The sensitivity of testing slurry samples for ESBL/AmpC herd status is less than 100%, especially for detecting herds with a low ESBL/AmpC prevalence. Therefore, whereas herds that tested positive for ESBL/AmpC-producing E. coli in slurry were defined as positive herds, herds with negative slurry samples were defined as unsuspected. Isolates of ESBL- and AmpC-producing E. coli were further characterized by detection and typing of their ESBL/AmpC gene. At the initial sampling, a comprehensive questionnaire was conducted at the participating farms. The farmers were asked questions about management practices potentially associated with the ESBL/AMPC herd status. Also, data on antimicrobial purchases during 2011 were acquired to evaluate whether the animal-defined daily dose of antimicrobials per year at farm level was associated with the ESBL/AmpC herd status. Multivariable logistic regression models were used to determine the association between management practices and the ESBL/AmpC herd status. Six months after the initial slurry sampling, 10 positive herds and 10 herds that had an unsuspected ESBL/AmpC herd status during the first visit were resampled. At each farm, slurry samples and feces from 24 individual cows were collected to evaluate within herd dynamics. During the first sampling, ESBL/AmpC-producing E. coli were isolated from the slurry samples collected at 41% of the herds. In total, 37 isolates were further characterized, revealing 7 different ESBL genes (blaCTX-M-1, -2, -14, -15, -32, -55 and blaTEM-52), 1 plasmid-encoded AmpC gene (blaCMY-2), and 1 chromosomally encoded ampC gene (ampC type 3). The total animal-defined daily dose of antimicrobials per year at farm level was not significantly different between ESBL/AmpC-positive and unsuspected dairy herds. The use of third- and fourth-generation cephalosporins, however, was found to be associated with ESBL/AmpC status, with higher use of these antimicrobials resulting in a significant higher odds to be ESBL/AmpC-positive. Management factors that were associated with a higher odds of being ESBL/AmpC-positive were treatment of all cases of clinical mastitis with antimicrobials, a higher proportion of calves treated with antimicrobials, not applying teat sealants in all cows at dry off, and the use of a floor scraper. This last association, however, was considered a methodological effect rather than a true risk factor. On 5 of the 10 initially positive farms, no ESBL/AmpC-producing E. coli were cultured from the slurry or any of the individual cow samples collected during the second sampling. In 4 of the initially unsuspected farms, slurry or individual cow samples tested positive during the second sampling. In conclusion, ESBL/AmpC could frequently be cultured from slurry samples collected from Dutch dairy farms and the ESBL/AmpC genes carried by the isolates were consistent with those reported earlier. The use of third- and fourth-generation cephalosporins appeared to be associated the ESBL/AmpC herd status.

Gonggrijp MA ，Santman-Berends IMGA ，Heuvelink AE ，Buter GJ ，van Schaik G ，Hage JJ ，Lam TJGM ... -  《-》

Prevalence of extended-spectrum and AmpC β-lactamase-producing Escherichia coli in Dutch dairy herds.

This study was conducted to assess: (1) a change in between-herd prevalence of extended-spectrum and AmpC β-lactamase-producing Escherichia coli (ESBL/AmpC-EC) between 2011 and 2013, the period during which the antimicrobial policy in animal husbandry in the Netherlands changed significantly, and (2) the prevalence of ESBL/AmpC-EC in individual calves, young stock, and dairy cows in the Netherlands. In 196 randomly selected conventional dairy herds, faecal samples were collected from calves (maximum n = 15), and randomly selected young stock (n = 5) and dairy cows (n = 15). Additionally, fresh faecal samples were collected from five different places on the floors where the dairy cows were housed. Samples were screened for E. coli with non-wild type susceptibility for cefotaxime and isolates were phenotypically confirmed as ESBL/AmpC-producing by disc diffusion, using cefotaxime and ceftazidime with and without clavulanic acid, and cefoxitin. Samples containing ESBL/AmpC-EC were examined semi-quantitatively. In 59.6% of the dairy herds one or more samples tested positive for ESBL/AmpC-EC. The between-herd prevalence based on floor samples in 2013 (18.0%) was significantly lower than the prevalence in 2011 based on comparable samples (32.7%). The individual animal prevalence of ESBL/AmpC-EC, with a minimum shedding level of 103 cfu/g of faeces, was 19.3% in calves, 0.9% in young stock, and 0.8% in dairy cows. Although ESBL/AmpC-EC was found in the majority of dairy herds, the herd prevalence declined significantly between 2011 and 2013. Calves were found to have both, a much higher individual animal prevalence and a higher level of shedding than young stock and cows.

Heuvelink AE ，Gonggrijp MA ，Buter RGJ ，Ter Bogt-Kappert CC ，van Schaik G ，Velthuis AGJ ，Lam TJGM ... -  《-》

Prevalence of extended-spectrum and AmpC β-lactamase-producing Escherichia coli in young calves on Dutch dairy farms.

In young calves on dairy farms the animal prevalence of extended-spectrum and AmpC β-lactamase-producing Escherichia coli (ESBL/AmpC-EC) is significantly higher compared with the animal prevalence in young stock and dairy cows. Hitherto it was unknown at what age antimicrobial resistant bacteria appear for the first time in the gut of calves on dairy farms, and how long these infections persist. The aim of this study was to examine the prevalence of ESBL/AmpC-EC, the number of excreted ESBL/AmpC-EC (in cfu/g of feces), as well as the ESBL/AmpC genotypes in young dairy calves (0-21 d of age) and the variation of these parameters between calves of different ages. Next to this, the course of shedding ESBL/AmpC-EC during the first year in dairy calves was studied. In a cross-sectional study, fecal samples from 748 calves, from 0 to 88 d of age, on 188 Dutch dairy farms were collected. The prevalence of calves testing positive for ESBL/AmpC-EC in a phenotypic assay was determined for different age categories (per 2 d of age). Positive samples were subjected to a semiquantitative test to determine the numbers of ESBL/AmpC-EC per gram of feces and for a selection of ESBL/AmpC-EC isolates the ESBL/AmpC genotype was determined. Ten of the 188 farms were selected for a longitudinal study based on the presence of at least 1 female calf with ESBL/Amp-EC in the cross-sectional study. These farms were additionally visited 3 times with a 4-mo interval. All calves that were sampled in the cross-sectional study were, if still present, resampled during the follow-up visits. Results show that from the day of birth ESBL/AmpC-EC can be present in the gut of calves. The phenotypic prevalence of ESBL/AmpC-EC was 33.3% in 0- to 21-d-old calves and 28.4% in 22- to 88-d-old calves. The prevalence of ESBL/AmpC-EC positive calves varied per age category among calves up to 21 d of age: significant increases and decreases at an early age were shown. Results of the longitudinal study show that after 4, 8, and 12 mo the prevalence of ESBL/AmpC-EC positive calves dropped to 3.8% (2/53), 5.8% (3/52), and 2.0% (1/49), respectively. This indicates that early gut colonization in young calves with ESBL/AmpC-EC is transient and does not lead to long-term shedding of these bacteria.

Gonggrijp MA ，Velthuis AGJ ，Heuvelink AE ，van den Heuvel KWH ，Ter Bogt-Kappert CC ，Buter GJ ，van Schaik G ，Lam TJGM ... -  《-》

Methicillin-resistant Staphylococcus aureus and extended-spectrum and AmpC β-lactamase-producing Escherichia coli in broilers and in people living and/or working on organic broiler farms.

The aim of this study was to estimate the prevalence of methicillin-resistant Staphylococcus aureus (MRSA) and extended-spectrum and AmpC β-lactamase (ESBL/AmpC)-producing Escherichia coli among broilers, and humans living and/or working on organic broiler farms; further characterise isolates; and compare these results with those from conventional farms. In the Netherlands, only 9 certified organic broiler farms were present. On 8 of these farms, 60 throat swabs and 20 cloacal swabs were taken per farm for MRSA and ESBL/AmpC-E. coli detection, respectively, at an average age of both 34 (T1) and 68 (T2) days. Faecal swabs and questionnaires were returned by 27 out of 36 humans. For selected ESBL/AmpC-producing E. coli isolates, phylogenetic groups, β-lactamase genes, plasmid families, and sequence types were determined. MRSA was not detected in broiler and human samples. ESBL/AmpC-producing E. coli were isolated from broilers on 7/8 farms at T1 and on all farms at T2. Furthermore, 3 farmers at T1, and 2 farmers and 1 family member at T2 were positive. Genes found in broilers and humans were almost exclusively blaCTX-M-1 and blaCMY-2. Given the high overall human ESBL/AmpC-prevalence (18.5%), which is similar to conventional farms, contact with live broilers is assumed a risk factor for carriage. Farm and sample-level prevalence at T1 are consistent with those from conventional farms. At T2, just before slaughter, sample-level prevalence of ESBL/AmpC-E. coli appears to have decreased (94.3% vs. 80%), which could have important consequences for contamination of retail meat.

Huijbers PM ，van Hoek AH ，Graat EA ，Haenen AP ，Florijn A ，Hengeveld PD ，van Duijkeren E ... -  《-》