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1.  Complete Genome Sequences of IncI1 Plasmids Carrying Extended-Spectrum β-Lactamase Genes 
Genome Announcements  2014;2(4):e00859-14.
Extended spectrum beta-lactamases (ESBLs) confer resistance to clinically relevant antibiotics. Often, the resistance genes are carried by conjugative plasmids which are responsible for dissemination. Five IncI1 plasmids carrying ESBLs from commensal and clinical Escherichia coli isolates were completely sequenced and annotated along with a non-ESBL carrying IncI1 plasmid.
PMCID: PMC4148731  PMID: 25169863
2.  Salmonella Subtypes with Increased MICs for Azithromycin in Travelers Returned to the Netherlands 
Emerging Infectious Diseases  2014;20(4):705-708.
Antimicrobial susceptibility was analyzed for 354 typhoidal Salmonella isolates collected during 1999–2012 in the Netherlands. In 16.1% of all isolates and in 23.8% of all isolates that showed increased MICs for ciprofloxacin, the MIC for azithromycin was increased. This resistance may complicate empirical treatment of enteric fever.
PMCID: PMC3966360  PMID: 24655478
Salmonella; typhi; paratyphi; typhoid; bacteria; enteric fever; cephalosporin; fluoroquinolone; azithromycin; drug resistance; travel medicine; the Netherlands
3.  The IncI1 plasmid carrying the blaCTX-M-1 gene persists in in vitro culture of a Escherichia coli strain from broilers 
BMC Microbiology  2014;14:77.
Commensal bacteria are a reservoir for antimicrobial-resistance genes. In the Netherlands, bacteria producing Extended Spectrum Beta-Lactamases (ESBL) are found on chicken-meat and in the gut of broilers at a high prevalence and the predominant ESBL-gene is the blaCTX-M-1 located on IncI1 plasmids. We aim to determine the fitness costs of this plasmid for the bacterium.
We investigated the conjugation dynamics of IncI1 plasmids carrying the blaCTX-M-1 gene in a batch culture and its impact on the population dynamics of three E. coli populations: donors, recipients and transconjugants. The intrinsic growth rate (ψ), maximum density (K) and lag-phase (λ) of the populations were estimated as well as the conjugation coefficient. Loss of the plasmid by transconjugants was either assumed constant or depended on the effective growth rate of the transconjugants.
Parameters were estimated from experiments with pure culture of donors, recipients and transconjugants and with mixed culture of donors and recipients with a duration of 24 or 48 hours. Extrapolation of the results was compared to a 3-months experiment in which a mixed culture of recipient and transconjugant was regularly diluted in new medium.
No differences in estimated growth parameters (ψ, K or λ) were found between donor, recipient and transconjugant, and plasmid loss was not observed. The conjugation coefficient of transconjugants was 104 times larger than that of the donor. In the 3-months experiment, the proportion of transconjugants did not decrease, indicating no or very small fitness costs.
In vitro the IncI1 plasmid carrying the blaCTX-M-1 gene imposes no or negligible fitness costs on its E. coli host, and persists without antimicrobial usage.
PMCID: PMC3987674  PMID: 24666793
ESBL; Antibiotic; Antimicrobial; Resistance; Poultry; Chicken; Livestock; Persistence; E. coli; Mathematical model
4.  Presence of ESBL/AmpC -Producing Escherichia coli in the Broiler Production Pyramid: A Descriptive Study 
PLoS ONE  2013;8(11):e79005.
Broilers and broiler meat products are highly contaminated with extended spectrum beta-lactamase (ESBL) or plasmid-mediated AmpC beta-lactamase producing Escherichia coli and are considered to be a source for human infections. Both horizontal and vertical transmission might play a role in the presence of these strains in broilers. As not much is known about the presence of these strains in the whole production pyramid, the epidemiology of ESBL/AmpC-producing E. coli in the Dutch broiler production pyramid was examined. Cloacal swabs of Grandparent stock (GPS) birds (one−/two-days (breed A and B), 18 and 31 weeks old (breed A)), one-day old Parent stock birds (breed A and B) and broiler chickens of increasing age (breed A) were selectively cultured to detect ESBL/AmpC-producing isolates. ESBL/AmpC-producing isolates were found at all levels in the broiler production pyramid in both broiler breeds examined. Prevalence was already relatively high at the top of the broiler production pyramid. At broiler farms ESBL/AmpC producing E. coli were still present in the environment of the poultry house after cleaning and disinfection. Feed samples taken in the poultry house also became contaminated with ESBL/AmpC producing E. coli after one or more production weeks. The prevalence of ESBL/AmpC-positive birds at broiler farms increased within the first week from 0–24% to 96–100% independent of the use of antibiotics and stayed 100% until slaughter. In GPS breed A, prevalence at 2 days, 18 weeks and 31 weeks stayed below 50% except when beta-lactam antibiotics were administered. In that case prevalence increased to 100%. Interventions minimizing ESBL/AmpC contamination in broilers should focus on preventing horizontal and vertical spread, especially in relation to broiler production farms.
PMCID: PMC3820706  PMID: 24244401
5.  Consumption of Antimicrobials in Pigs, Veal Calves, and Broilers in The Netherlands: Quantitative Results of Nationwide Collection of Data in 2011 
PLoS ONE  2013;8(10):e77525.
In 2011, Dutch animal production sectors started recording veterinary antimicrobial consumption. These data are used by the Netherlands Veterinary Medicines Authority to create transparency in and define benchmark indicators for veterinary consumption of antimicrobials. This paper presents the results of sector wide consumption of antimicrobials, in the form of prescriptions or deliveries, for all pig, veal calf, and broiler farms. Data were used to calculate animal defined daily dosages per year (ADDD/Y) per pig or veal calf farm. For broiler farms, number of animal treatment days per year was calculated. Furthermore, data were used to calculate the consumption of specific antimicrobial classes per administration route per pig or veal calf farm. The distribution of antimicrobial consumption per farm varied greatly within and between farm categories. All categories, except for rosé starter farms, showed a highly right skewed distribution with a long tail. Median ADDD/Y values varied from 1.2 ADDD/Y for rosé finisher farms to 83.2 ADDD/Y for rosé starter farms, with 28.6 ADDD/Y for white veal calf farms. Median consumption in pig farms was 9.3 ADDD/Y for production pig farms and 3.0 ADDD/Y for slaughter pig farms. Median consumption in broiler farms was 20.9 ATD/Y. Regarding specific antimicrobial classes, fluoroquinolones were mainly used on veal calf farms, but in low quantities: P75 range was 0 – 0.99 ADDD/Y, and 0 – 0.04 ADDD/Y in pig farms. The P75 range for 3rd/4th-generation cephalosporins was 0 – 0.07 ADDD/Y for veal calf farms, and 0 – 0.1 ADDD/Y for pig farms. The insights obtained from these results, and the full transparency obtained by monitoring antimicrobial consumption per farm, will help reduce antimicrobial consumption and endorse antimicrobial stewardship. The wide and skewed distribution in consumption has important practical and methodological implications for benchmarking, surveillance and future analysis of trends.
PMCID: PMC3804574  PMID: 24204857
6.  Prevalence of antimicrobial resistance among bacterial pathogens isolated from cattle in different European countries: 2002–2004 
The project "Antibiotic resistance in bacteria of animal origin – II" (ARBAO-II) was funded by the European Union (FAIR5-QLK2-2002-01146) for the period 2003–2005, with the aim to establish a continuous monitoring of antimicrobial susceptibility among veterinary laboratories in European countries based on validated and harmonised methodologies. Available summary data of the susceptibility testing of the bacterial pathogens from the different laboratories were collected.
Antimicrobial susceptibility data for several bovine pathogens were obtained over a three year period (2002–2004). Each year the participating laboratories were requested to fill in excel-file templates with national summary data on the occurrence of antimicrobial resistance from different bacterial species.
A proficiency test (EQAS – external quality assurance system) for antimicrobial susceptibility testing was conducted each year to test the accuracy of antimicrobial susceptibility testing in the participating laboratories. The data from this testing demonstrated that for the species included in the EQAS the results are comparable between countries.
Data from 25,241 isolates were collected from 13 European countries. For Staphylococcus aureus from bovine mastitis major differences were apparent in the occurrence of resistance between countries and between the different antimicrobial agents tested. The highest frequency of resistance was observed for penicillin. For Mannheimia haemolytica resistance to ampicillin, tetracycline and trimethoprim/sulphonamide were observed in France, the Netherlands and Portugal. All isolates of Pasteurella multocida isolated in Finland and most of those from Denmark, England (and Wales), Italy and Sweden were susceptible to the majority of the antimicrobials. Streptococcus dysgalactiae and Streptococcus uberis isolates from Sweden were fully susceptible. For the other countries some resistance was observed to tetracycline, gentamicin and erythromycin. More resistance and variation of the resistance levels between countries were observed for Escherichia coli compared to the other bacterial species investigated.
In general, isolates from Denmark, England (and Wales), the Netherlands, Norway, Sweden and Switzerland showed low frequencies of resistance, whereas many isolates from Belgium, France, Italy, Latvia and Spain were resistant to most antimicrobials tested. In the future, data on the prevalence of resistance should be used to develop guidelines for appropriate antimicrobial use in veterinary medicine.
PMCID: PMC2486267  PMID: 18611246
7.  Occurrence of antimicrobial resistance among bacterial pathogens and indicator bacteria in pigs in different European countries from year 2002 – 2004: the ARBAO-II study 
The project "Antibiotic resistance in bacteria of animal origin – II" (ARBAO-II) was funded by the European Union (FAIR5-QLK2-2002-01146) for the period 2003–05. The aim of this project was to establish a program for the continuous monitoring of antimicrobial susceptibility of pathogenic and indicator bacteria from food animals using validated and harmonised methodologies. In this report the first data on the occurrence of antimicrobial resistance among bacteria causing infections in pigs are reported.
Susceptibility data from 17,642 isolates of pathogens and indicator bacteria including Actinobacillus pleuropneumoniae, Streptococcus suis and Escherichia coli isolated from pigs were collected from fifteen European countries in 2002–2004.
Data for A. pleuropneumoniae from infected pigs were submitted from five countries. Most of the isolates from Denmark were susceptible to all drugs tested with the exceptions of a low frequency of resistance to tetracycline and trimethoprim – sulphonamide.
Data for S. suis were obtained from six countries. In general, a high level of resistance to tetracycline (48.0 – 92.0%) and erythromycin (29.1 – 75.0%) was observed in all countries whereas the level of resistance to ciprofloxacin and penicillin differed between the reporting countries. Isolates from England (and Wales), France and The Netherlands were all susceptible to penicillin. In contrast the proportion of strains resistant to ciprofloxacin ranged from 12.6 to 79.0% (2004) and to penicillin from 8.1 – 13.0% (2004) in Poland and Portugal.
Data for E. coli from infected and healthy pigs were obtained from eleven countries. The data reveal a high level of resistance to tetracyclines, streptomycin and ampicillin among infected pigs whereas in healthy pigs the frequency of resistance was lower.
Bacterial resistance to some antimicrobials was frequent with different levels of resistance being observed to several antimicrobial agents in different countries. The occurrence of resistance varied distinctly between isolates from healthy and diseased pigs, with the isolates from healthy pigs generally showing a lower level of resistance than those from diseased pigs.
The study suggests that the choice of antimicrobials used for the treatment of diseased animals should preferably be based on knowledge of the local pattern of resistance.
PMCID: PMC2459172  PMID: 18554407
8.  Salmonella Gene rma (ramA) and Multiple-Drug-Resistant Salmonella enterica Serovar Typhimurium 
MarA and its homologue, RamA, have been implicated in multidrug resistance (MDR). RamA overexpression in Salmonella enterica serovar Typhimurium and Escherichia coli conferred MDR independently of marA. Inactivation of ramA did not affect the antibiotic susceptibilities of wild-type S. enterica serovar Typhimurium or 15 unrelated clinical MDR isolates. Thus, ramA overexpression is not a common MDR mechanism in Salmonella.
PMCID: PMC415616  PMID: 15155237
9.  Molecular Diversity and Evolutionary Relationships of Tn1546-Like Elements in Enterococci from Humans and Animals 
We report on a detailed study on the molecular diversity and evolutionary relationships of Tn1546-like elements in vancomycin-resistant enterococci (VRE) from humans and animals. Restriction fragment length polymorphism (RFLP) analysis of the VanA transposon of 97 VRE revealed seven different Tn1546 types. Subsequent sequencing of the complete VanA transposons of 13 VRE isolates representing the seven RFLP types followed by sequencing of the identified polymorphic regions in 84 other VanA transposons resulted in the identification of 22 different Tn1546 derivatives. Differences between the Tn1546 types included point mutations in orf1, vanS, vanA, vanX, and vanY. Moreover, insertions of an IS1216V-IS3-like element in orf1, of IS1251 in the vanS-vanH intergenic region, and of IS1216V in the vanX-vanY intergenic region were found. The presence of insertion sequence elements was often associated with deletions in Tn1546. Identical Tn1546 types were found among isolates from humans and farm animals in The Netherlands, suggesting the sharing of a common vancomycin resistance gene pool. Application of the genetic analysis of Tn1546 to VRE isolates causing infections in hospitals in Oxford, United Kingdom, and Chicago, Ill., suggested the possibility of the horizontal transmission of the vancomycin resistance transposon. The genetic diversity in Tn1546 combined with epidemiological data suggest that the DNA polymorphism among Tn1546 variants can successfully be exploited for the tracing of the routes of transmission of vancomycin resistance genes.
PMCID: PMC89148  PMID: 10049255

Results 1-9 (9)