Meeting on antimicrobial resistance in animal health · Meeting on antimicrobial resistance . in animal health . ... emergence can lead to therapeutic impasses that have dramatic

18 November 2011

Meeting on antimicrobial resistance in animal health

Veterinary public health and antimicrobials:

ANSES’s work

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Veterinary public health and antimicrobials: ANSES’s work

In recent years, the emergence of antimicrobial-resistant bacteria, both in humans and in animals, has become a major concern in public and animal health. Some of these cases of emergence can lead to therapeutic impasses that have dramatic consequences in the treatment of certain serious infections. The methods currently available to scientists are unable to assess the extent to which the use of antimicrobials in animals is responsible for the emergence and selection of resistance genes in human bacteria, as the pathogens involved in the spread of resistance genes are frequently common to humans and animals. It is therefore necessary to promote the rational use of antimicrobials, in human medicine as in veterinary medicine, in order to preserve their effectiveness in both humans and animals.

Since 1999, at the request of the French Ministry of Agriculture, ANSES has worked to define and monitor relevant indicators for antimicrobial consumption in animal production sectors and domestic pets, as well as to measure changes in antimicrobial resistance of bacteria found in animals. In line with this, the Agency publishes each year a report entitled "Sales survey of Veterinary Medicinal Products containing Antimicrobials in France" and a report from its surveillance network for antimicrobial resistance in pathogenic bacteria of animal origin (RESAPATH), as well as data on the monitoring of zoonotic indicator bacteria obtained through these reference missions.

The perspective offered by these reports, supplemented by consolidated data from specific animal sector surveys (poultry in 2008, pigs in 2009, cattle and rabbits in 2010) illustrates the efforts made in this area by the sectors, and enables a better assessment to be made of their impact on the emergence of resistant bacteria. In recent years, ANSES has proved to be particularly vigilant about the emergence of resistance to fluoroquinolones and third-generation cephalosporins (3GC), antimicrobials of critical importance in the treatment of certain serious infections in humans. Although there was concern about the increase in resistance to 3GC in the pig sector between 2006 and 2009, the policy of a moratorium on their use as a preventive measure, imposed by this sector at the end of 2010, should begin bearing fruit in 2011. The moderate increase in the number of strains resistant to both 3GC and fluoroquinolones is an early encouraging result that should be confirmed when examining strains isolated in 2011, after a full year of the above moratorium. In contrast, the results recorded in 2010 for resistance to 3GC in the hen/chicken sector are disturbing, since the percentage of resistant E. coli, isolated from infections in these animals, has doubled every year since 2006, even though this resistance has stabilised at a low level in the turkey sector. The situation with regard to fluoroquinolone resistance in the hen/chicken and turkey sectors did however seem to have stabilised in 2010. This dramatic increase in 3GC

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resistance in E. coli isolated in the hen/chicken sector should quickly lead to questions being asked about the conditions of use of these compounds, and a debate on the risks of spreading resistant E. coli within production pyramids.

In connection with its mission to assess the risks associated with the emergence of antimicrobial-resistant bacteria, ANSES recently completed a call for experts in order to set up a working group to respond to the solicited request on antimicrobial resistance entitled "Risks of emergence of antimicrobial resistance associated with patterns of antimicrobial use in the field of animal health". This group will assess the risk of emergence of antimicrobial resistance, but will also propose alternative solutions that will inform animal health stakeholders of possible ways to reduce the use of antimicrobials.

As part of its mission to provide scientific support to the French Ministry of Agriculture, ANSES participates in the work of the National Committee for rational use of antibiotics in veterinary medicine, which proposed an action plan in May 2011 that resulted in the national plan to reduce the risk of antimicrobial resistance, launched by the Minister of Agriculture.

In addition, as part of its research missions, ANSES is continuing its work on the molecular mechanisms of antimicrobial resistance, its biological cost for bacteria, pharmaco-epidemiology, monitoring sales, the flow of genes and resistant bacteria in the environment, as well as the influence of treatment practices (including the injection of 3GC in the embryonated egg or one-day chick) on the emergence of resistance. Besides this work, which is directly related to monitoring and understanding emergence, the basic work of the research teams is driven by the desire to improve animal health by taking into account all its components. Our epidemiology teams are therefore seeking to define the risk factors of disease expression, whether related to herd management, configuration of buildings, prophylactic treatment practices, environmental factors, human factors, etc. Our laboratory bacteriologists, in turn, are developing and consolidating faster and safer techniques for detecting pathogens and their resistance to antimicrobials, and methods for assessing overall herd health, as well as undertaking work on new vaccine valences, and other such factors that contribute to improved diagnosis and prevention.

Finally, our teams in the experimental departments are helping, both through work in their organisations and in the field, to define the factors of disease expression on single- and multi-microbial models that are also leading to a better understanding of pathogen expression and association.

Moreover, ANSES is continuing and reinforcing its work at EU level, in particular through its driving role in the establishment of a European surveillance scheme for antimicrobial use (European Surveillance of Antimicrobial Consumption - ESVAC) led by EMA (European Medicines Agency), or its action within the HMA (Heads of Medicines Agencies) group, for the implementation of the European action plan on antimicrobial resistance adopted in February 2011.

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All this work will ultimately contribute to a better understanding of the disease “condition" and of prevention, which remains the best way of avoiding the use of antimicrobials, which are nevertheless irreplaceable for the treatment of certain complex bacterial diseases. For this reason, their rational use is a challenge that must be met in order to preserve their effectiveness in both public and animal health.

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Veterinary public health and antimicrobials:

ANSES’s work

1. Gaining a better understanding of uses and practices 5 1.1. Monitoring sales of veterinary antimicrobials in 2010 1.2. Understanding use practices in farming

- Study on antimicrobials uses in veterinary medicine in rabbits - Studies on antimicrobials uses in veterinary medicine in ruminants

2. Monitoring the presence of resistance in bacteria 11 2.1. An example of surveillance of antimicrobial resistance in bacteria that are pathogenic for animals 2.2: The importance of surveillance of resistance in commensal bacteria

3. Risk assessment: 14 Solicited request entitled “Risks of emergence of antimicrobial resistance associated with patterns of antimicrobial use in the field of animal health”

4. Factors for development of antimicrobial resistance: the research themes completed at ANSES 16

5. The French Agency for Veterinary Medicinal Products: an animal health authority 20

6. Antimicrobial resistance: the ANSES laboratories involved 21

7. Annexes 24

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1. Gaining a better understanding of uses and practices

The study of the use of antimicrobials in veterinary medicine in France is based on the complementary nature of two schemes, firstly the annual monitoring of sales of veterinary medicinal products containing antimicrobials and secondly, specific studies conducted in farms or with veterinary practitioners.

1.1 Monitoring sales of veterinary antimicrobials in 2010 Since 1999, the Agency has been monitoring annual sales of veterinary antimicrobials. Based on the OIE guideline on "Monitoring the quantities of antimicrobials used in animal husbandry", this monitoring is carried out in collaboration with the French Union for the Veterinary Medicinal Product and Reagent Industry (SIMV) and is funded by the Ministry of Agriculture.

Why should sales of antimicrobials be monitored? The information gathered is one of the essential elements, together with monitoring of bacterial resistance, needed for assessing the risks associated with antimicrobial resistance and proposing measures to manage these risks and monitor changes in practices so as to evaluate their effectiveness. How is this monitoring conducted? Monitoring of antimicrobial sales is based on the annual declaration of antimicrobial sales by the laboratories marketing them. These data can be combined with other sources of information (reported turnover, prescription surveys, etc.). The information collected from the laboratories covers 100% of authorised drugs1

How should these results be interpreted?

. For the second consecutive year, the laboratories reported, for most drugs, the breakdown by target species. For some species, these declarations were confirmed by surveys of veterinary prescription and use in farming.

The tonnages of antimicrobials sold do not precisely reflect their use because of differences in activity and dose between different antimicrobials. Recent antimicrobials are generally more potent and require the administration of a smaller dose. To assess animal consumption or exposure to antimicrobials, it is necessary to consider, in particular, the dosage and duration of administration, but also changes in the animal population over time. Thus, a decrease in sales volume does not necessarily mean a decrease in the rate of use.

1 The extra-label use of veterinary medicinal products is partly taken into consideration in the manufacturers’ declarations. Exceptional prescription and extra-label use of human drugs or extemporaneous preparations containing antibiotics under the provisions of the cascade approach (Article L. 5143-4 of the French Code of Public Health) is not taken into account.

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What are the main changes observed?

- Sales volumes In 2010, total sales of antimicrobials amounted to 1014 tonnes. This was the lowest tonnage recorded since monitoring began. The 2010 results confirm the fall in volumes seen in previous years. Compared to 1999, the tonnage of antimicrobials sold in 2010 thus fell by 23%. Compared to data from the previous year, sales decreased by 3.6%.

- Animal exposure to antimicrobials For monitoring animal exposure, the most relevant indicator is the animal level of exposure to antimicrobials (ALEA), because it takes into account information on treatment (dose and duration) and potential users (mass of the animal population potentially consuming the antimicrobials). Between 1999 and 2010, the exposure of animals to antimicrobials administered via oral and parenteral routes, all classes combined, increased by 12.4%. Three phases seem to have emerged: an increase in exposure from 1999 to 2007, a decrease in exposure in 2008 and 2009, and finally, stable exposure between 2009 and 2010.

However, this overall trend must be qualified according to the target species and classes of compounds.

Are there any developments that justify the implementation of specific measures? Third- and fourth-generation cephalosporins and fluoroquinolones are considered as particularly important in human medicine because they are among the only alternatives for the treatment of certain infectious diseases in humans. These two classes of compound have been available in veterinary medicine for the past fifteen years. According to European recommendations, these antimicrobials should be reserved for second-line therapeutic treatment. Over twelve years of monitoring, animals’ exposure to fluoroquinolones has almost doubled, while exposure to cephalosporins has almost tripled. Over the last three years of monitoring, exposure to these two classes of antimicrobials, however, has stabilised:

- Exposure to fluoroquinolones decreased slightly in 2010 compared to 2009 (-0.24%). - Exposure to cephalosporins decreased in 2009 then increased again in 2010. In 2010, the level of exposure was equivalent to that of 2008.

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Changes in exposure to Quinolones and

Fluoroquinolones

Changes in exposure to Cephalosporins

0

0,005

0,01

0,015

0,02

0,025

1999

2000

2001

2002

2003

2004

2005

2006

2007

2008

2009

2010

Céphalosporines 3&4G

Céphalosporines 1&2G

The curves showing the changes in animal exposure to these two antimicrobial classes clearly show a trend towards stabilisation after a period of strong growth. It is important to confirm this trend, which is likely related to the communication campaigns on the use of these two classes of compounds. In late 2010 and early 2011, several announcements were made by professionals and ministries (the moratorium on the use of cephalosporins in pigs reported by the professional organisations, the target to reduce consumption by 25% over 5 years set by the Minister of Agriculture). In addition, the National Committee for rational use of antibiotics in veterinary medicine proposed a draft action plan in May 2011 which will shortly be submitted to the Minister of Agriculture. Despite the stabilisation observed since 2008, the sharp increase in sales and exposure to fluoroquinolones and cephalosporins since 1999 is disturbing. Data collected over the next few years will be used to assess the impact of measures carried out in France in terms of rational use of antimicrobials. What is the situation in France compared to other European countries? At present, few European countries have a system for monitoring sales of antimicrobials. The European Medicines Agency (EMA) recently published a retrospective pilot study (in which France actively participated) of the years 2005-2009 for nine European countries. The published data relate to sales rather than exposure and cannot be used to make a comparison between countries in terms of antimicrobial consumption. Irrespective of the situation in France compared to other European countries, efforts must continue to rationalise and minimise the use of antimicrobials as far as possible. This use should be reserved for cases where it is essential to animal health and welfare.

Cephalosporins 3&4G

Cephalosporins 1&2G

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1.2 Understanding use practices in farming

Studies in farms have been conducted by the ANSES laboratories. They are usually targeted at a particular sector and based on limited data samples (geographical area, study duration). The aim of these studies is generally to explore the variability of a measure (such as prescribed dosages), to investigate the farming factors associated with variability of uses, or to compare the properties of different methods for estimating antimicrobial consumption.

- Example of a study in the rabbit sector by the ANSES Ploufragan-Plouzané Laboratory

Knowledge of antimicrobial use practices is an essential prerequisite for preparing a debate on future trends. At the joint request of the rabbit farming sector and the French Agency for Veterinary Medicinal Products, the ANSES Ploufragan-Plouzané Laboratory conducted a survey in 2010 and 2011 on the use of antimicrobials in organised meat rabbit farming. The aim of the study was to obtain a precise and comprehensive initial picture of the antimicrobials used and the circumstances of their use, and to explore the variability in uses between farms so as to identify the associated factors. This was a qualitative and quantitative, descriptive and analytical study, whose implementation required a representative sample of farms to be established from which reliable, accurate and informative data could be collected. Use of antimicrobials was then quantified from inventories of purchases in 2009, the circumstances of their use were documented from the treatment history of one batch per farm, and changes in use were explored through the updating of the purchase inventory for 2010. The random selection of the farms, then their inclusion in the study and the collection of data required and received the cooperation of all the professionals from the sector. This resulted in a high participation rate (81%) and led to a nationally representative sample being obtained of 113 breeding/fattening farms.

Antimicrobials used and routes of administration This study established that rabbit farms mainly use compounds belonging to the classes of tetracyclines, aminoglycosides and peptides. The critical antimicrobial classes of fluoroquinolones, cephalosporins and macrolides accounted for the lowest amounts and the least frequent occurrences, with a relative share of less than 10% of all antimicrobials, either purchased or used. The overall estimates obtained are quantitatively similar to those documented by the ANMV in its national monitoring of sales of veterinary medicines containing antimicrobials. Some isolated qualitative differences were observed, such as a discrepancy in the use of sulfonamides, which was quantitatively lower in the study sample.

The main route of administration is oral, via food and then drinking water. Injections represent a minority and are almost exclusively reserved for breeding stock. Although only 40% of purchases were intended for female rabbits, they had greater exposure to antimicrobials than meat rabbits. Digestive diseases are the principal reasons for antimicrobial use, but a

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distinction can be made between female rabbits, mainly affected by respiratory infections, and male rabbits, primarily subject to digestive disorders.

Discussion

The study of the variability in the quantities purchased revealed significant differences between farms. The farm distribution curve follows a normal distribution, with most farms having "average" use and few farms that could be described as "high" or "low" users of antimicrobials. There has therefore been a change in practices through a collective industry approach and commitment, which seems to have been initiated by the professionals themselves. Comparing 2009 and 2010 does indeed show a decrease, from 10 to 15% depending on the indicator used, in the quantities of antimicrobials purchased. This decrease is especially apparent for administration via medicated feed.

- Example of studies in the ruminants sector by the ANSES Lyon Laboratory Monitoring the consumption of antimicrobials is an essential element in the fight against antimicrobial resistance. To complement the data generated by the French Agency for Veterinary Medicinal Products (ANMV) in its monitoring of antimicrobial sales, ANSES carries out pharmaco-epidemiology studies to characterise antimicrobial use in the various animal production sectors, under real farming conditions. The objective is to describe antimicrobial utilisation, i.e., to identify the main antimicrobials used and the conditions of their use. Qualitative surveys were conducted through questionnaires on the two most recent antimicrobial treatments prescribed by veterinarians or implemented by farmers in the cattle (2007), sheep (2007), and goat (in progress) sectors. In the cattle (excluding veal calves) and sheep sectors, the vast majority of antimicrobial treatments are individual. In dairy sheep or cattle, the most common pathological context behind the use of antimicrobials relates to udder conditions (primarily treated by the farmer), followed by musculoskeletal problems in cattle. Veterinarians in the cattle sector mainly prescribe antimicrobials for obstetric and gynaecological problems and respiratory diseases. Antimicrobials used The observation unit used here is the antimicrobial treatment itself, with each antimicrobial class being assigned a value proportional to its use in relation to all the prescribed or administered treatments. As some treatments may include several commercial antimicrobials, which themselves may contain several compounds from different antimicrobial classes, the totals can exceed 100%. The antimicrobials used vary depending on the pathological context, but those used most widely, irrespective of the sector, are penicillins (40 to 60% of treatments), aminoglycosides (40 to 50% of treatments) and tetracyclines (14% for veterinarians in the cattle sector, 30% for cattle and sheep farmers). In the cattle sector, cephalosporins and fluoroquinolones are used in respectively 19% and 24% of treatments and are more frequent in milk production

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than in meat production. However, these two classes are only rarely used in the sheep sector. Medical context Only 48% and 27% of treatments carried out respectively by cattle and sheep farmers involve a veterinary consultation. Respecting the MAs In the cattle industry, apart from a few anecdotal cases, all the antimicrobials used have been granted marketing authorisation (MA) for the bovine species. Using them for a pathological indication other than those mentioned in the MA (i.e. extra-label use for pathology) accounts for 13% of veterinary prescriptions and 7% of treatments by farmers. Conversely, in the sheep sector, as the therapeutic arsenal is more limited, use of antimicrobials outside the conditions specified in the MA for the species is more common (16%), although extra-label use with respect to the therapeutic indication is similar (8%) to that observed in the cattle sector. Analysis of dosage (dose, frequency and duration of administration) prescribed by veterinarians in the cattle sector indicates that 70% of prescriptions comply with the indications of the MA, 14% are higher and 16% lower than those defined in the MA.

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2. Monitoring the presence of resistance in bacteria Because it provides data on trends and emerging resistance in bacteria found in animals, surveillance is both a decision support tool for responsible prescribing by veterinarians and an information tool for managers. Over the past decade, the Agency has significantly strengthened its surveillance scheme and played a very active role in building the European surveillance approach.

At European level, the Zoonoses Directive2 specifically recommends surveillance of antimicrobial resistance in Salmonella and Campylobacter, and in indicator bacteria3

ANSES’s work on surveillance of bacterial resistance is based on the activities of several complementary systems and on field studies.

. EFSA has drawn up technical recommendations for this surveillance, and a network of European and national reference laboratories coordinated by the ANSES Fougères Laboratory, which is also a European Union reference laboratory, is working to harmonise the techniques used and improve the quality of the analyses.

- RESAPATH4

- The

collects data on antimicrobial resistance in bacteria isolated from sick animals during veterinary diagnosis.

Salmonella5

- The annual surveillance plans implemented by the French Directorate General of Food, in collaboration with the Agency’s laboratories as part of their NRL mission, organises the collection of faeces or caeca from healthy animals at the slaughterhouse, from which are isolated sentinel

Network collects non-human Salmonella strains (isolated from food, the environment or animal production) for serotyping and study of their sensitivity to antimicrobials.

6 or zoonotic7

Based on this work, the Agency publishes a FARM

bacterial strains in the cattle, pig and poultry sectors.

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report every two years, summarising this information. The last edition was published in 2010.

2.1. An example of surveillance of antimicrobial resistance in bacteria that are pathogenic for animals The RESAPATH network, established in 1982 under the name RESABO (for BOvine), is now structured around sixty analytical laboratories that collect antibiogram data on pathogenic animal bacteria from ninety-four French départements. These are isolated from samples from sick animals treated by veterinary practitioners as part of their routine customer business. Under ANSES’s governance, RESAPATH is run jointly by the Lyon and Ploufragan-Plouzané

2 Directive 2003/99/EC of the European Parliament and of the Council of 17 November 2003 on the monitoring of zoonoses and zoonotic agents 3 E. coli, Enterococcus faecium 4 Coordinated by the Lyon and Ploufragan laboratories 5 Coordinated by the Maisons-Alfort Laboratory for Food Safety 6 E. coli, Enterococcus faecium 7 Campylobacter spp, and several Salmonella isolates 8 French Antimicrobial Resistance Monitoring in bacteria of animal origin

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laboratories, and many measures (including the organisation of annual inter-laboratory trials) improve the accuracy of the data. In 2010, a total of 24,274 antibiograms were recorded, giving the network a level of coverage equivalent to that of the largest French medical networks on the subject. RESAPATH is also the only veterinary network to be a member of the French National Observatory for Epidemiology of Bacterial Resistance to Antimicrobials (ONERBA), which brings together a further sixteen surveillance networks for bacterial resistance in humans, in general practice and in hospitals. This integration enables the permanent sharing of human and animal data on bacterial resistance, and ensures a joint vision, which is particularly important at a time when efforts to reduce rates of resistance need to be unified. As well as the cattle, and later the pig and poultry sectors, which were historically central to surveillance concerns, RESAPATH has recently expanded its scope to cover all animal species, including pets. The surveillance results therefore allow the Agency to identify priority areas, which are also correlated with the results of epidemiological studies on uses and reflect the contribution of each animal production sector to this issue. Obviously, considering overall antimicrobial resistance in animals is not a satisfactory basis for recommending relevant levers for action, and it is essential to continue to highlight these aspects in sufficient detail. Furthermore, in addition to analysing the raw data on resistance by animal sector/pathology/bacterial species, RESAPATH’s phenotypic monitoring is very closely associated - this is a key point - with laboratory molecular surveillance, which can assess more precisely the scope of public health issues when identical resistance mechanisms or bacterial clones are found in humans and animals (extended-spectrum beta-lactamases (ESBLs), Escherichia coli ST131, MRSA ST398, etc.). This characterisation by genotype also provides keys to understanding the epidemiology of animal resistance itself, for example on assumptions about the spread of resistance between animal sectors by highlighting identical molecular media. This same approach is also followed specifically for Salmonella through the Salmonella Network’s collection of isolates, and helps identify the emergence of new antimicrobial resistance phenotypes in the different animal production sectors and the related mechanisms (ESBLs, CMY-2). Finally, these surveillance data, by identifying the key difficulties in animal resistance, are a natural starting point for the development of recommendations for use in veterinary antimicrobial therapy. They can then be compared and contrasted with all the components of the treatment action, primarily feedback from practitioners on the clinical effectiveness of the compounds, the presence or absence of alternatives to antimicrobial treatment, the zoonotic constraints specific to each sector and the current therapeutic indications from manufacturers for each of their commercial products, including known extra-label uses. 2.2 The importance of surveillance of resistance in commensal bacteria European Regulations9

9 According to Directive 2003/99/EC

stipulate that Member States shall collect, assess and report their data on zoonoses (animal diseases transmissible to humans), the agents responsible

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(zoonotic agents), antimicrobial resistance and outbreaks of foodborne illness. Surveillance of resistance in commensal bacteria (E. coli and Enterococcus sp.) shall be done on a voluntary basis. Based on the analysis of faeces or caeca collected randomly at the slaughterhouse from major food-producing species (pigs, broiler chickens, turkey, cattle), such surveillance provides a useful indicator for monitoring changes in antimicrobial resistance as a function of antimicrobial use. Based on a randomly sampled isolate, it provides an indicator of resistance in the dominant commensal flora. Having a standard method for determining the minimum inhibitory concentration in the context of our reference missions facilitates the inter-comparison of data at European level. Surveillance of resistance in commensal flora, which has continued for over ten years, has been used to measure the effect of phasing out the use of growth factors in the 2000s on the frequencies of resistance to vancomycin, virginiamycin or avilamycin in strains of Enterococcus faecium. Thus the percentage of resistance to avilamycin fell from 70% in 2000 to less than 5% in 2008 for strains of E. faecium isolated from broilers, while streptogramin resistance fell from 50% to less than 5% for strains isolated from chickens, and from 35% to 10% for those isolated from pigs. For E. coli strains isolated from chickens, there was a reduction in rates of resistance to several aminoglycosides. This surveillance reveals the effects of changes in use in veterinary antimicrobial therapy and in particular the emergence of resistance to cephalosporins and fluoroquinolones in broiler chickens. The same samples can also be used to assess the prevalence of certain resistance phenotypes in commensal flora by obtaining isolates using selective media. Several European Union Member States undertake this surveillance in their major production sectors and provide EFSA with comparable data that can be analysed in terms of changing distributions of minimum inhibitory concentrations and percentages of resistance, on the basis of harmonised interpretation. In coming years, increased reporting to EFSA of antibiograms by strains will improve the surveillance of multiple resistance in commensal species in the European Union. Surveillance of resistance in indicator bacteria via random isolation is a useful tool for assessing trends and obtaining information on the risk of consumer exposure from contaminated food. Coupled with prevalence studies using selective media, which are more complex to implement, it helps to assess the presence in sub-dominant flora of resistance phenotypes that may emerge under selective pressure.

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3. Risk assessment: Solicited request entitled “Risks of emergence of antimicrobial resistance associated with patterns of antimicrobial use in the field of animal health”

Background For the past ten years the Agency has worked on the theme of antimicrobial resistance. In particular, it produced a report in 2006 entitled "Veterinary use of antibiotics, bacterial resistance and consequences for human health". This report describes in particular the mechanisms of bacterial resistance that may result from the use of antimicrobials in animals, and the mechanisms of dissemination of antimicrobial resistance in bacteria that are important in human medicine. Today, there is mobilisation around the issue of antimicrobial resistance at all levels in governments, public health and veterinary agencies, as well as among professionals. In order to coordinate and maximise all the stakeholders’ efforts, the French Ministries of Agriculture and Health, together with ANSES, have set up a National Coordinating Committee for rational use of antimicrobials in veterinary medicine. The Ministry of Agriculture has also launched a national plan to reduce the risks of antimicrobial resistance in veterinary medicine. In this context, ANSES decided to deploy its study, research and risk assessment resources to formulate, on an independent scientific basis, recommendations on preventing the development of antimicrobial resistance in the field of animal health. To do this, it set up an expert group to conduct an assessment of the risks of emergence and dissemination of antimicrobial resistance associated with patterns of antimicrobial use in the animal health field (livestock and domestic pets). The working group’s roadmap

- A first step will involve listing the uses of antimicrobials in the different farming sectors and in domestic pets as well as the tools for monitoring antimicrobial resistance in bacteria isolated from animals, relying in particular on the work carried out over the past few years by the ANSES laboratories.

- Subsequently, an assessment will be conducted of the risk of emergence and dissemination of antimicrobial resistance in the different sectors. It will identify the unsafe practices leading to the selection of multi-resistant bacteria, and measure, according to their indications, the effects of prophylactic, curative or metaphylactic use of antimicrobials in veterinary medicine for livestock and pets.

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- Finally, proposals will be made to validate treatment regimens that reduce or eliminate unsafe practices in veterinary medicine or to suggest some alternative solutions to antimicrobials in farming that will nonetheless maintain control of animal health.

To form the working group, ANSES issued a call for applications from March to July 2011. Twenty-nine experts were selected, all combining field and laboratory skills, or research expertise, in the fields of microbiology, animal pathology, animal science, pharmacology and pharmaco-epidemiology. The following animal groups or sectors were targeted: ruminants, pigs, poultry, rabbits, fish, horses and domestic cats. Because of the working group’s extensive mandate, its mission does not include consideration of the environment as a reservoir of resistant bacteria and resistance genes. Nor is it intended to address the risk of human exposure to bacteria of animal origin carrying antimicrobial resistance genes, regardless of the exposure route, direct or indirect, via food or the environment. The working group will have a predominant "animal" focus. Progress of work After a phase of identification of the available literature and existing inventories on the use of antimicrobials in veterinary medicine, the working group will meet in plenary session for its first meeting on 21 November 2011. Sub-groups will be formed by animal sector, for the first stage of compiling practices of antimicrobial use. This is important because it will determine the quality of the risk assessment and the recommendations that follow. Various means are used to achieve a sufficiently detailed understanding of the practices for the different species. The sub-groups may use the tools already developed by the ANSES laboratories and supplement their information with interviews with professionals. This compilation should be mostly completed by mid-2012. The working group will also be able to interview the various stakeholders affected by this issue. Completion of the group’s work is scheduled for mid-2013 with an opinion being issued in the autumn of 2013.

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4. Factors for development of antimicrobial resistance: the research themes completed at ANSES

The fight to maintain the effectiveness of antimicrobials must necessarily be based on a research effort, as was outlined in the National Plans to preserve the effectiveness of antimicrobials in human and veterinary medicine (led respectively by the DGS and the DGAL). The processes responsible for selection of antimicrobial resistance in a bacterial population, emergence of a new mechanism of resistance, and spread of resistance genes or resistant bacteria are still poorly understood, whether in pathogenic or commensal flora. However, an analysis of the consequences of exposure to antimicrobials in human and animal populations on bacterial resistance to these compounds is essential for effective risk management.

Discussions are ongoing in the scientific communities for both human health and animal health to define research needs more precisely. The questions are indeed common because there is no segregation between bacterial populations of environmental10

The ANSES laboratories, in addition to their reference mandate for the Ministry of Agriculture and based on their surveillance activity, conduct research on antimicrobial resistance. Four of them have worked on this theme for many years: Fougères, Ploufragan, Lyon and Maisons-Alfort. Their work is carried out alone or in partnership with external agencies (Institut Pasteur, INSERM, INRA, Cemagref, National Veterinary Schools and Universities). In addition, the French Agency for Veterinary Medicinal Products monitors sales of antimicrobials in veterinary medicine.

, human and animal origin, from the perspective of exchanges of DNA, a natural phenomenon which is one of the mechanisms behind the evolution of living things.

Research priorities for animal and public health at ANSES:

Theme 1

As part of their epidemiological surveillance mission, some laboratories are studying the prevalence of antimicrobial resistance in certain zoonotic bacteria or specific pathogens. They can therefore detect the emergence of antimicrobial-resistant strains in animals, food and the environment. In connection with this mission, research teams are working to characterise resistance genes and interactions between mechanisms of resistance and the metabolic function of the bacteria.

: reinforce research on the descriptive epidemiology of bacterial zoonoses and changes in their agents’ susceptibility/resistance to antimicrobials

10 The main natural source of plasmid resistance genes disseminated in pathogenic bacteria

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Theme 2

• Analyse sales of antimicrobials in veterinary medicine in order to quantify their use in the different sectors. The ANMV is conducting this research with the epidemiology teams at Ploufragan and Lyon.

: analyse the characteristics of antimicrobial use in animals

• Characterise the use of antimicrobials in rural veterinary medicine in each sector according to the modes of treatment: prevention, early treatment and conventional curative treatment. The Lyon and Ploufragan laboratories are conducting pharmaco-epidemiology studies ranging from multi-year monitoring of antimicrobial use in the various livestock sectors (pigs, poultry and ruminants) to the analysis of prescribing and dispensing practices for veterinary use antimicrobials.

• Characterise the use of antimicrobials in canine and feline veterinary medicine.

Theme 3

• Impact of antimicrobial use on the emergence and persistence of bacterial resistance in animals

: study the impact of antimicrobial use on bacterial resistance especially in the intestinal flora

The epidemiological studies in poultry include (i) the association in chickens between carriage of resistant bacteria at the slaughterhouse and the exposure of batches sampled during their life, and (ii) measurement of the time between changes in use and impact on resistance through the joint study of two chronological series, one of practices and the other of resistance.

The pharmacological and biological studies, also in poultry, include (i) assessing the risk/benefit of the use of fluoroquinolones, and (ii) the biological cost of antimicrobial resistance in Campylobacter. Other studies focus on ruminants (E. coli and beta-lactams; Campylobacter and fluoroquinolones) or pigs (the study of selection conditions in the intestine during treatment, mathematical modelling of the emergence and spread of resistant bacteria in farms). Finally, ongoing studies are evaluating the link between antimicrobial resistance and virulence of bacteria.

In July 2011, ANSES initiated a research project to assess the impact on the intestinal flora of poultry of treatment practices involving third-generation cephalosporins in hatcheries.

• Increased risk of transmission of antimicrobial resistance in human populations treated with antimicrobials.

This issue is being addressed by the Fougères laboratory in collaboration with INRA/ENV Toulouse using a "rodent" simulation model (simulating both the animal and the human): this model is used to study the effects of exposure to antimicrobial treatments (or antimicrobial residues) on:

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o intestinal barrier flora and the risk of introduction of resistant pathogenic bacteria.

o the risk of transmission of plasmid resistance genes in a simplified model to bacteria in unbalanced intestinal flora.

Theme 4: study the spread of bacterial resistance from animals to humans by direct11 or indirect12

The Ploufragan and Fougères laboratories have initiated multi-partner research on the spread of antimicrobial resistance in the environment, via the ANR project "Evaluation of the use of fluoroquinolones in poultry", a study of the persistence or selection of fluoroquinolone-resistant bacteria in manure, or in soil and water after spreading; and a study of gene transfer to soil bacteria. The results of this major research programme will be presented during the meeting on antimicrobial resistance to be held by ANSES on 18 November 2011.

exposure

Several strategies have been used to assess the benefits and risks associated with the use of fluoroquinolones in poultry production. For example, an original model of avian colibacillosis was developed and used to 1) compare the effectiveness of several antimicrobials including fluoroquinolones, 2) obtain data regarding the fate of the antimicrobial in the body, 3) assess the impact of treatments on the resistance of the digestive flora and 4) determine in a microcosm study the persistence of antimicrobials and resistant bacteria following spreading of manure from treated birds. The environmental consequences were also analysed in "full-scale" trials after storage or spreading on experimental plots in order to determine the concentrations of antimicrobials in manure, soil or environmental water and to screen for the presence of resistant bacteria in the soil several months after spreading. Finally, epidemiological surveys have been conducted and an economic model was constructed to evaluate the comparative economic benefit of the use of fluoroquinolones in the fight against colibacillosis and to specify the impact of pathological episodes on the health quality of carcasses. The most significant results of this programme include the limited persistence over time of ciprofloxacin-resistant enterobacteria, which account for about 60% of total enterobacteria seven days after spreading of manure from treated animals, but which decrease over time and disappear a month after spreading.

These four themes form the basis for future development of a quantitative risk analysis and the direction of risk management measures. The research needs for the fight against the development of antimicrobial resistance are considerable and ANSES will be helping to specify them clearly by animal production sector as part of the future plan for a rational use of antimicrobials in veterinary medicine.

11 At work for professionals in animal husbandry, at home for pet owners 12 Via food of animal origin, or via the environment (soil and water) contaminated by effluents from farms and processing industries

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Veterinary pharmacovigilance involves monitoring the risk of adverse reactions caused by veterinary medicinal products. The ANMV collects information on: • the effectiveness of drugs; • the consequences of drug treatments in terms of residues in foodstuffs of animal origin (validity of the withdrawal period*); • their impact on the environment. It also takes into account uses that do not comply with the instructions and labelling of veterinary medicinal products.

* Minimum time between administration of a drug and marketing of foodstuffs of animal origin.

5. The French Agency for Veterinary Medicinal Products: an animal health authority ANSES is responsible for overseeing the application of regulations regarding veterinary medicinal products in the spheres of competence attributed by the French Public Health Code. Its mission includes scientific assessment and full control and decision-making powers. It exercises this specific authority via the French Agency for Veterinary Medicinal Products (ANMV). Active in several international authorities, it participates in discussions concerning veterinary pharmacy and contributes to the drafting of numerous regulatory and technical documents in this field. In particular it is a collaborating centre of the World Organisation for Animal Health (OIE), and participates in the work of the Food and Agriculture Organization (FAO) of the United Nations and the World Health Organization (WHO). Finally it is a member of the European Heads of Medicines Agencies (HMA). The ANMV assesses • national and European marketing authorisation dossiers for veterinary medicinal products, • European dossiers on maximum residue limits of veterinary medicinal products in foodstuffs of animal origin. It authorises • placing on the market of medicinal products, • their clinical trials, • the opening of pharmaceutical establishments for exploitation, manufacture, wholesale distribution, export and import of veterinary medicinal products:

- import of veterinary medicinal products; - provisional use of medicinal products authorised elsewhere in Europe or in third countries.

It monitors • the risk of adverse effects of drugs, • the quality of drugs, • the operation of veterinary pharmaceutical establishments, • advertising of veterinary medicinal products, • imports and exports of veterinary medicinal products.

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6. Antimicrobial resistance: the ANSES laboratories involved

- Fougères The Fougères laboratory, with 65 staff members, mainly contributes to improving knowledge of: • the benefits and risks of veterinary medicinal product and disinfectant use by the food processing industry; • the risks associated with food contaminants. Its work focuses on: • screening for residues of veterinary medicinal products in foodstuffs of animal origin; • the antimicrobial effectiveness of antimicrobials and disinfectants, and resistance to these products; • assessing the genetic toxicity of food contaminants. As the reference laboratory for veterinary medicinal product residues and antimicrobial resistance, it is involved in the surveillance and control of these health risks. Because it participates in the standardisation of methods for determining the effectiveness of disinfectants, it also contributes to the assessment of biocidal products in Europe. Data from its work contribute to the proper use of veterinary medicinal products and antimicrobial hygiene products in France and Europe. Through its toxicology research work, it provides the data necessary for the risk assessment of contaminants and residues in food.

- Lyon The Lyon Laboratory, with 75 staff members, mainly contributes to improving the health of ruminants. It specialises in transmissible spongiform encephalopathies (TSEs), antimicrobial resistance, mycoplasmoses and certain viral diseases (such as Rift Valley fever): • it studies the agents responsible for major and emerging diseases • it studies pathogenic bacteria in ruminants (resistance to antimicrobials and virulence) • it monitors the emergence and spread of diseases and analyses their causes • as the National Reference Laboratory (NRL) for several diseases, it provides scientific and technical support for veterinary inspections carried out by the authorities (validation of data and analytical methods, training of field laboratories, management of epidemiological surveillance data, etc.). The Lyon Laboratory also has a unit dedicated to monitoring and studying the resistance of plant bioaggressors to phytosanitary products.

- Maisons-Alfort - Laboratory for Food Safety This laboratory, with 150 staff members, studies the biological and physico-chemical hazards that can affect food safety.

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In its area of expertise, it helps accomplish the Agency’s reference, research, surveillance, epidemiology and scientific and technical expertise missions. The Laboratory provides scientific support to public decision makers, both general expertise on the quality and safety of foodstuffs, and more specific expertise on certain processing industries (milk and dairy products in particular). It is involved in identifying, monitoring and analysing the risks associated with major food hazards: • Microbiological hazards linked to raw materials, conditions of production and preparation of foodstuffs; • Physico-chemical hazards in the environment or generated by agro-industrial processes, which can be found in food. As part of its antimicrobial resistance reference missions, it is involved in determining antibiograms (COFRAC accreditation) and minimum inhibitory concentrations, and in molecular identification of resistance mechanisms in strains of Salmonella and Staphylococcus aureus. These data are used by the Salmonella Network and the DGAL surveillance plans. They are regularly communicated through bulletins and reports from the Salmonella Network, the FARM report, and through joint ANSES/DGAL/DGS epidemiological bulletins. Research on molecular mechanisms of resistance is also conducted and published in scientific journals.

- Ploufragan-Plouzané The laboratory has 200 staff members at its two sites in Brittany. It specialises in poultry, rabbit, pig and fish farming. It contributes to improving animal health and welfare as well as to the quality of food of animal origin. Its main research programmes focus on: • adapting statistical methods to epidemiology • antimicrobial resistance and pharmaco-epidemiology • poultry and pig welfare • calicivirus in rabbits and hares • Campylobacter, Salmonella, Listeria, Yersinia • fish immunology and immunotoxicology • the impact of environmental pollutants on animals • avian influenza, Gumboro disease and avian metapneumovirus • influenza/influenza-like illnesses in pigs • digestive flora/pathogenic bacteria interactions • host/pathogen interactions • modelling of infectious phenomena • porcine respiratory disease • poultry parasitology

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• viral diseases in fish • swine fevers • vaccinology/vectorology • emerging diseases

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Annex:

ANSES publications relating to antimicrobial resistance

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Publications relating to antimicrobial resistance

2006 Publications

2006 to 2011

Butaye, P., Michael, G.B., Schwarz, S., Barrett, T.J., Brisabois, A., White, D.G. 2006. The clonal spread of multidrug-resistant non-typhi Salmonella serotypes. Microbes Infect 8, 1891-1897.

Cailhol, J., Lailler, R., Bouvet, P., La Vieille, S., Gauchard, F., Sanders, P., Brisabois, A. 2006. Trends in antimicrobial resistance phenotypes in non-typhoid Salmonellae from human and poultry origins in France. Epidemiol. Infect. 134, 171-178.

Cardinale E, Rose V, Perrier-Gros-Claude JD, Tall F, Rivoal K, Mead GC, Salvat G. 2006. Genetic characterisation and antimicrobial resistance of Campylobacter spp. Isolated from poultry and human in Senegal. J. Appl. Microbiol. 100 (1): 209-214.

Hasman H, Kempf I, Chidaine B, Cariolet R, Ersboll A K, Houe H, Bruun Hansen H C, Aarestrup Fm, 2006. Copper resistance in Enterococcus faecium, mediated by the tcrB gene, is selected by supplementation of pig feed with copper sulfate. Appl. Environ. Microbiol. 72, 5784-5789.

Kehrenberg C., Meunier D., Targant H., Cloeckaert A., Schwarz S., Madec J.-Y. (2006) Plasmid-mediated florfenicol resistance in Pasteurella trehalosi. Journal of Antimicrobial Agents and Chemotherapy, 58, 1, 13-17.

Le Carrou J, Laurentie M, Kobisch M, Gautier-Bouchardon AV, 2006. Persistence of Mycoplasma hyopneumoniae in experimentally infected pigs after a marbofloxacin treatment, and detection of mutations in the parC gene. Antimicrob. Agents Chemother. 50: 1959-1966.

Le Carrou J, Reinhardt Ak, Kempf I, Gautier-Bouchardon AV, 2006. Persistence of Mycoplasma synoviae in hens after two enrofloxacin treatments and detection of mutations in the parC gene. Vet. Res. 37, 1-24.

Meunier D, Jouy E, Lazizzera C, Kobisch M and Madec JY, 2006. CTX-M-1 and CTX-M-15 type beta-lactamases in clinical Escherichia coli isolates recovered from food-producing animals in France. International Journal of Antimicrobial Agents, 28, 402-407. .

Moore JE, Barton MD, Blair IS, Corcoran D, Dooley JSG, Fanning S, Kempf I, Lastovica AJ, Lowery CJ, Matsuda M, Mcdowell DA, Mcmahon A, Millar BC, Rao JR, Rooney PJ, Seal BS, Snelling WJ, Tolba O, 2006. The epidemiology of antimicrobial resistance in Campylobacter. Microbes and Infection Emerging Antimicrobial Resistance Mechanisms in Zoonotic Food-borne pathogens Salmonella and Campylobacter, in Microbes and Infection. 8, 1955-66.

Pringle M, Aarestrup F, Bergsjo B, Fossi M, Jouy E, Landen A, Mevius D, Perry K, Teale C, Thomson J, Skrzypczak T, Veldman K And Franklin A, 2006. Quality control ranges for antimicrobial susceptibility testing by broth dilution of the Brachyspira hyodysenteriae type strain (ATCC 27164). Microbial Drug Resistance, 12, 219-221.

Stevens, A., Kaboré, Y., Perrier-Gros-Claude, J.D., Millemann, Y., Brisabois, A., Catteau, M., Calvin, J.F., Dufour, B. 2006. Prevalence and antibiotic-resistance of Salmonella isolated from beef sampled from the slaughterhouse and from retailers in Dakar. Int J Food Microbiol 110, 178-186.

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2007 Publications

Chauvin, C., C. Clement, M. Bruneau, and D. Pommeret. 2007. Time-patterns of antibiotic exposure in poultry production-A Markov chains exploratory study of nature and consequences. Preventive Veterinary Medicine 80:230-240.

Gallay, A., Prouzet-Mauleon, V., Kempf, I., Lehours, P., Labadi, L., Camou, C., Denis, M., De Valk, H., Desenclos, J.-C., and Megraud, F. (2007). Campylobacter antimicrobial drug resistance among humans, broiler chickens, and pigs, France. Emerg Infect Dis 13, 259-266.

Girlich, D., Poirel, L., Carattoli, A., Kempf, I., Lartigue, M.-F., Bertini, A., and Nordmann, P. (2007). Extended-spectrum b-lactamase CTX-M-1 in Escherichia coli isolates from healthy poultry in France. Appl Environ Microbiol 73, 4681-4685.

Kayser, E., Morignat, E., Meunier, D., Madec, J. Y., Calavas, D., and Botrel, M. A. (2007). Investigation statistique des multirésistances aux antibiotiques chez des souches d'Escherichia coli isolées chez les bovins de 2002 à 2005. Epidémiologie et santé animale 52, 59-74.

Kempf, I. (2007). L'antibiorésistance de Campylobacter en filières avicole et porcine. Le Nouveau Praticien Vétérinaire - Elevages et santé 5, 59-62. 2008 Publications

Aarestrup, F., Battisti, A., Bengtsson, B., Piriz Duran, S., Emborg, H.-D., Kahlmeter, G., Mevius, D., Regula, G., Sanders, P., Teale, C., et al. (2008). Harmonised monitoring of antimicrobial resistance in Salmonella and Campylobacter isolates from food animals in the European Union. Clin Microbiol Infect 14, 522-533.

Chauvin, C., Querrec, M., Perot, A., Guillemot, D., and Sanders, P. (2008). Impact of antimicrobial drug usage measures on the identification of heavy users, patterns of usage of the different antimicrobial classes and time-trends evolution. J Vet Pharmacol Ther 31, 301-311.

Doublet, B., Douard, G., Targant, H., Meunier, D., Madec, J.-Y., and Cloeckaert, A. (2008). Antibiotic marker modifications of lambda Red and FLP helper plasmids, pKD46 and pCP20, for inactivation of chromosomal genes using PCR products in multidrug-resistant strains. J Microbiol Methods 75, 359-361.

Egorova, S., Timinouni, M., Demartin, M., Granier, S. A., Whichard, J. M., Sangal, V., Fabre, L., Delauné, A., Pardos, M., Millemann, Y., et al. (2008). Ceftriaxone-resistant Salmonella enterica serotype Newport, France. Emerg Infect Dis 14, 954-957.

Hendriksen, R. S., Mevius, D. J., Schroeter, A., Teale, C., Jouy, E., Butaye, P., Franco, A., Utinane, A., Amado, A., Moreno, M., et al. (2008). Occurrence of antimicrobial resistance among bacterial pathogens and indicator bacteria in pigs in different European countries from year 2002 - 2004: The ARBAO-II study. Acta Vet Scand 50.

Hendriksen, R. S., Mevius, D. J., Schroeter, A., Teale, C., Meunier, D., Butaye, P., Franco, A., Utinane, A., Amado, A., Moreno, M., et al. (2008). Prevalence of antimicrobial resistance among bacterial pathogens isolated from cattle in different European countries: 2002-2004. Acta Vet Scand 50. Kempf, I., Hellard, G., Perrin-Guyomard, A., Gicquel-Bruneau, M., Sanders, P., and Leclercq, R. (2008). Prevalence of high-level vancomycin-resistant enterococci in French broilers and pigs. Int J Antimicrob Agents 32, 463-464.

Madec, J.-Y., Lazizzera, C., Chatre, P., Meunier, D., Martin, S., Lepage, G., Menard, M.-F., Lebreton, P., and Rambaud, T. (2008). Prevalence of fecal carriage of acquired expanded-spectrum

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cephalosporin resistance in Enterobacteriaceae strains from cattle in France. J Clin Microbiol 46, 1566-1567. Moulin G., Cavalie P., Pellanne I., Chevance A., Laval A., Millemann Y., Colin P., Chauvin C., on behalf of the 'Antimicrobial Resistance' ad hoc Group of the French Food Safety Agency, 2008. A comparison of antimicrobial usage in human and veterinary medicine in France from 1999 to 2005. Journal of Antimicrobial Chemotherapy, 62, 617-625. Peyrat, M. B., C. Soumet, P. Maris, and P. Sanders. 2008. Phenotypes and genotypes of campylobacter strains isolated after cleaning and disinfection in poultry slaughterhouses. Veterinary Microbiology 128:313-326. Peyrat, M. B., C. Soumet, P. Maris, and P. Sanders. 2008. Recovery of Campylobacter jejuni from surfaces of poultry slaughterhouses after cleaning and disinfection procedures: Analysis of a potential source of carcass contamination. International Journal of Food Microbiology 124:188-194. Vachée A., Varon E., Jouy E., Meunier D. ; pour le conseil scientifique de l’Onerba. 2008. Sensibilité aux antibiotiques chez les streptocoques et les entérocoques: données de l'Onerba. Pathologie Biologie 57:240-244.

2009 Publications Arpin, C., Quentin, C., Grobost, F., Cambau, E., Robert, J., Dubois, V., Coulange, L., and André, C. (2009). Nationwide survey of extended-spectrum β-lactamase-producing Enterobacteriaceae in the French community setting. J Antimicrob Chemother 63, 1205-1214. Botrel M.-A., Morignat E., Meunier D., Madec J.-Y. and Calavas D. (2009) Identifying antimicrobial multiresistance patterns of Escherichia coli sampled from diarrhoeic calves by cluster analysis techniques: a way to guide research on multiresistance mechanisms. Zoonoses and Public Health. 1-7 Chauvin, C., 2009, Impact of generic introduction on antimicrobial usages - A time-series analysis. J. Vet. Pharmacol. Ther. 32, 111-112 Chazel, M., Jouy, E., Meunier, D., Haenni, M., Gay, E., Calavas, D., and Madec, J. Y. (2009). RESAPATH: réseau d'épidémiosurveillance de l'antibiorésistance des bactéries pathogènes animales. Bulletin épidémiologique 34, 7-8. Denis, M., Chidaine, B., Laisney, M.J., Kempf, I., Rivoal, K., Megraud, F., Fravalo, P., 2009. Comparison of genetic profiles of Campylobacter strains isolated from poultry, pig and Campylobacter human infections in Brittany, France. Pathol Biol (Paris) 57, 23-29.

Doublet, B., Granier, S. A., Robin, F., Bonnet, R., Fabre, L., Brisabois, A., Cloeckaert, A., and Weill, F.-X. (2009). Novel plasmid-encoded ceftazidime-hydrolyzing CTX-M-53 extended-spectrum -lactamase from Salmonella enterica serotypes Westhampton and Senftenberg. Antimicrob Agents Chemother 53, 1944-1951.

Elgroud, R., Zerdoumi, F., Benazzouz, M., Bouzitouna-Bentchouala, C., Granier, S.A., Fremy, S., Brisabois, A., Dufour, B., Millemann, Y. 2009. Characteristics of Salmonella contamination of broilers and slaughterhouses in the region of Constantine (Algeria). Zoonoses and Public Health 56, 84-93.

Faure, S., Perrin-Guyomard, A., Delmas, J. M., and Laurentie, M. (2009). Impact of therapeutic treatment with β-lactam on transfer of the blaCTX-M-9 resistance gene from Salmonella enterica serovar Virchow to Escherichia coli in gnotobiotic rats. Appl Environ Microbiol 75, 5523-5528.

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Ferran, A. A., Kesteman, A. S., Toutain, P. L., and Bousquet-Mélou, A. (2009). Pharmacokinetic/pharmacodynamic analysis of the influence of inoculum size on the selection of resistance in Escherichia coli by a quinolone in a mouse thigh bacterial infection model. Antimicrob Agents Chemother 53, 3384-3390.

Greko, C. (2009). Reflection paper on the use of third and fourth generation cephalosporins in food producing animals in the European Union: Development of resistance and impact on human and animal health. J Vet Pharmacol Ther 32, 515-533.

Guillouzouic, A., Caroff, N., Dauvergne, S., Lepelletier, D., Perrin Guyomard, A., Kempf, I., Reynaud, A., and Corvec, S. (2009). MLST typing of Escherichia coli isolates overproducing AmpC β-lactamase. J Antimicrob Chemother 63, 1290-1292.

Haenni, M., Saras, E., Chĝtre, P., Meunier, D., Martin, S., Lepage, G., Ménard, M. F., Lebreton, P., Rambaud, T., and Madec, J. Y. (2009). VanA in Enterococcus faecium, Enterococcus faecalis, and Enterococcus casseliflavus detected in French cattle. Foodborne Pathog Dis 6, 1107-1111.

Kempf I, Zeitouni S. [The cost of antimicrobial resistance: Analysis and consequences.]. Pathol Biol (Paris). 2009 Nov 24.

Kesteman, A. S., Ferran, A. A., Perrin-Guyomard, A., Laurentie, M., Sanders, P., Toutain, P. L., and Bousquet-Melou, A. (2009). Influence of inoculum size and dosing regimen on the selection of marbofloxacin resistant mutants in experimental rat Klebsiella pneumoniae lung infection. J Vet Pharmacol Ther 32, 63.

Kesteman, A. S., Ferran, A. A., Perrin-Guyomard, A., Laurentie, M., Sanders, P., Toutain, P. L., and Bousquet-Mélou, A. (2009). Influence of inoculum size and marbofloxacin plasma exposure on the amplification of resistant subpopulations of Klebsiella pneumoniae in a rat lung infection model. Antimicrob Agents Chemother 53, 4740-4748.

Madec, J. Y. (2009). Les méthodes d'évaluation de la sensibilité des bactéries aux antibiotiques. Bulletin des GTV 49, 49-54.

Maurer, C., Meunier, D., and Madec, J.-Y. (2009). Shiga toxin Stx2 production is promoted by enrofloxacin in experimental in vitro-selected mutants of Escherichia coli O157:H7 resistant to fluoroquinolones. Foodborne Pathog Dis 6, 257-259. Meunier D., Jouy E., Lazizzera C., Doublet B., Kobisch M., Cloeckaert A. and Madec J.-Y. 2010. Plasmid-borne florfenicol and ceftiofur resistance encoded by the floR and blaCMY-2 genes in Escherichia coli isolates from diseased cattle in France. Journal of Medical Microbiology, 59:467-471.

Sanders, P. (2009). Use of fluoroquinolones and development of resistance in France and Europe. J Vet Pharmacol Ther 32, 52-53.

2010 Publications Botrel M.-A., Haenni M., Morignat E., Sulpice P., Madec J.-Y. and Calavas D.. (2010) Distribution and Antimicrobial Resistance of Clinical and Subclinical Mastitis Pathogens in Dairy Cows in Rhône-Alpes, France. Foodborne Pathogens and Diseases, 7 (5): 479-487.

Châtre P., Haenni M., Meunier D., Botrel M.-A., Calavas D. and Madec J.-Y.. (2010) Prevalence and Antimicrobial Resistance of Campylobacter jejuni and Campylobacter coli Isolated from Healthy Cattle between 2002 and 2006 in France. Journal of Food Protection, 73 (5): 825-831.

Cloeckaert, A., Praud, K., Lefevre, M., Doublet, B., Pardos, M., Granier, S.A., Brisabois, A., Weill, F.X. 2010. IncI1 plasmid carrying extended-spectrum-beta-lactamase gene blaCTX-M-1 in Salmonella

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enterica isolates from poultry and humans in France, 2003 to 2008. Antimicrob Agents Chemother 54, 4484-4486.

Haenni M., L. Galofaro, M. Giddey, M. Ythier, P. Majcherczyk, P. Moreillon and J.-Y. Madec (2010). Penicillin-binding protein (PBP) genes in Streptococcus uberis presenting a decreased susceptibility to penicillin. Antimicrobial Agents and Chemotherapy, 54 (3): 1140-1145

Haenni M., Saras E. and Madec J.-Y. (2010) Demonstration of a shift towards penicillin G resistance in the Streptococcus uberis population. The Journal of Microbiological Methods, 59 (Pt 8): 993-995

Haenni M., Targant H., Forest K., Sévin C., Tapprest J., Laugier C. and Madec J.-Y. (2010) Retrospective study of necropsy-associated coagulase-positive staphylococci in horses. Journal of Veterinary Diagnostic Investigation, 22 (6): 953-956.

Haenni M., Saras E., Bertin S., Leblond P., Madec J.-Y. and Payot S. (2010). Diversity and mobility of Integrative Conjugative Elements and link with antibioresistance in strains of S. agalactiae, S. dysgalactiae and S. uberis isolated from clinical mastitis in dairy cows. Applied and Environmental Microbiology, 76 (24): 7957-7965.

Hocquet D., Plésiat P., Dehecq B., Mariotte P., Talon D., Bertrand X., conseil scientifique de l’ONERBA. (2010) Nationwide investigation of extended-spectrum beta-lactamases, metallo-beta-lactamases, and extended-spectrum oxacillinases produced by ceftazidime-resistant Pseudomonas aeruginosa strains in France. Antimicrobial Agents Chemotherapy. 54 (8):3512-5.

Hopkins, K.L., Kirchner, M., Guerra, B., Granier, S.A., Lucarelli, C., Porrero, M.C., Jakubczak, A., Threlfall, E.J., Mevius, D.J. 2010. Multiresistant Salmonella enterica serovar 4,[5],12:i:- in Europe: a new pandemic strain? Euro Surveill 15, 19580.

Huehn, S., La Ragione, R.M., Anjum, M., Saunders, M., Woodward, M.J., Bunge, C., Helmuth, R., Hauser, E., Guerra, B., Beutlich, J., Brisabois, A., Peters, T., Svensson, L., Madajczak, G., Litrup, E., Imre, A., Herrera-Leon, S., Mevius, D., Newell, D.G., Malorny, B. 2010. Virulotyping and antimicrobial resistance typing of Salmonella enterica serovars relevant to human health in Europe. Foodborne Pathog Dis 7, 523-535.

Kempf, I., Zeitouni, S., sous presse. The cost of antimicrobial resistance: Analysis and consequences. Coût biologique de la résistance aux antibiotiques: analyse et conséquences. Pathologie Biologie (sous presse)

Meunier D., E. Jouy, C. Lazizzera, B. Doublet, M. Kobish, A. Cloeckaert and J.-Y. Madec. (2010) Plasmid-borne florfenicol and ceftiofur resistance encoded by the floR and CMY-2 genes in Escherichia coli isolates from diseased cattle in France. Journal of Medical Microbiology, 59: 467-471.

Le Devendec, L., Bouder, A., Dheilly, A., Hellard, G., Kempf, I. Persistence and spread of qnr, ESBL and ampC resistance genes in the digestive tract of chickens. Microbial Drug Resistance (accepted)

Catry, B., E. Van Duijkeren, M. C. Pomba, C. Greko, M. A. Moreno, S. Pyorala¤, M. Ruzauskas, P. Sanders, E. J. Threlfall, F. Ungemach, K. Torrneke, C. Munoz-Madero, and J. Torren-Edo. 2010. Reflection paper on MRSA in food-producing and companion animals: Epidemiology and control options for human and animal health. Epidemiology and Infection 138:626-644.

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Cerf, O., B. Carpentier, and P. Sanders. 2010. Tests for determining in-use concentrations of antibiotics and disinfectants are based on entirely different concepts: "Resistance" has different meanings. International Journal of Food Microbiology 136:247-254. Faure, S., A. Perrin-Guyomard, J. M. Delmas, P. Chatre, and M. Laurentie. 2010. Transfer of plasmid-mediated CTX-M-9 from Salmonella enterica serotype virchow to Enterobacteriaceae in human flora-associated rats treated with cefixime. Antimicrobial Agents and Chemotherapy 54:164-169. Kesteman, A. S., A. Perrin-Guyomard, M. Laurentie, P. Sanders, P. L. Toutain, and A. Bousquet-Mélou. 2010. Emergence of resistant Klebsiella pneumoniae in the intestinal tract during successful treatment of Klebsiella pneumoniae lung infection in rats. Antimicrobial Agents and Chemotherapy 54:2960-2964. Ruimy, R., Brisabois, A., Bernede, C., Skurnik, D., Barnat, S., Arlet, G., Momcilovic, S., Elbaz, S., Moury, F., Vibet, M.A., Courvalin, P., Guillemot, D., Andremont, A., 2010, Organic and conventional fruits and vegetables contain equivalent counts of Gram-negative bacteria expressing resistance to antibacterial agents. Environ Microbiol 12, 608-615. Sanders, P. 2010. Antimicrobial resistance of zoonotic bacteria: Current strategies in veterinary medicine. Résistance aux antibiotiques chez les bactéries d'origine animale: Actions en cours dans le secteur vétérinaire 26:930-935. Targant H., Ponsin C., Brunet C., Doublet B., Cloeckaert A., Madec J.-Y. and Meunier D.. (2010) Multidrug resistance in Salmonella enterica serotype Typhimurium isolated from diseased cattle from 2002 to 2007.. Foodborne Pathogens and Disease, 7 (4): 419-425. Targant H., Doublet B., Aarestrup F.A., Cloeckaert A. and Madec J.-Y. (2010) IS6100-mediated genetic rearrangement within the complex class 1 integron In104 of the Salmonella genomic island 1 (2010). Journal of Antimicrobial Chemotherapy, 65 (7): 1543-1545.

2011 Publications

Andraud M, Chauvin C, Sanders P, Laurentie M., 2011. Pharmacodynamic modelling of in vitro activity of marbofloxacin against E. coli strains. Antimicrob Agents Chemother, 55, 756-761.

Andraud M., Rose N., Laurentie M., Sanders P., Le Roux A., Cariolet R., Chauvin C., Jouy E. 2011. Estimation of transmission parameters of a fluoroquinolone-resistant Escherichia coli strain between pigs in experimental conditions. Veterinary Research, 42:44.

Bouzidi, N., Aoun, L., Dekhil, M., Granier, S.A., Poirel, L., Brisabois, A., Nordmann, P., Millemann, Y. 2011. Co-occurrence of aminoglycoside resistance gene armA in non-Typhi Salmonella isolates producing CTX-M-15 in Algeria. Journal of Antimicrobial Chemotherapy 66, 2180-2181. Bouzidi, N., Aoun, L., Zeghdoudi, M., Bensouilah, M., Elgroud, R., Oucief, I., Granier, S.A., Brisabois, A., Desquillet, L., Millemann, Y. 2011. Salmonella contamination of laying-hen flocks in two regions of Algeria. Food Research International, epub ahead of print Jun 12. Bugarel, M., Granier, S.A., Bonin, E., Vignaud, M.L., Roussel, S., Fach, P., Brisabois, A. 2011. Genetic diversity in monophasic (1,4,[5],12:i:- and 1,4,[5],12:-:1,2) and in non-motile (1,4,[5],12:-:-) variants of Salmonella enterica S. Typhimurium. Food Research International, epub ahead of print Jul 12.

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Bugarel, M., Granier, S.A., Weill, F.X., Fach, P., Brisabois, A. 2011. A multiplex real-time PCR assay targeting virulence and resistance genes in Salmonella enterica serotype Typhimurium. BMC Microbiology 11, 151. Dheilly, A., Bouder, A., Le Devendec, L., Hellard, G., Kempf, I., 2011, Clinical and microbial efficacy of antimicrobial treatments of experimental avian colibacillosis. Vet Microbiol 149, 422-429. Dheilly, A., Le Devendec, L., Mourand, G., Bouder, A., Jouy, E., Kempf , I., in press, Resistance gene transfer during treatments of experimental avian colibacillosis. Antimicrobial Agents and Chemotherapy (in press). Garcia-Migura, L., Sunde, M., Karlsmose, S., Veldman, K., Schroeter, A., Guerra, B., Granier, S.A., Perrin-Guyomard, A., Gicquel-Bruneau, M., Franco, A., Englund, S., Teale, C., Heiska, H., Clemente, L., Boerlin, P., Moreno, M.A., Daignault, D., Mevius, D., Hendriksen, R.S., Aarestrup, F.M. 2011. Establishing Streptomycin Epidemiological Cut-Off Values for Salmonella and Escherichia coli. Microbial Drug Resistance, epub ahead of print Jul 12. Granier, S.A., Hidalgo, L., San Millan, A., Escudero, J.A., Gutierrez, B., Brisabois, A., Gonzalez-Zorn, B. 2011. ArmA Methyltransferase in a Monophasic Salmonella enterica Isolate from Food. Antimicrobial Agents and Chemotherapy 55, 5262-5266. Granier, S.A., Moubareck, C., Colaneri, C., Lemire, A., Roussel, S., Dao, T.T., Courvalin, P., Brisabois, A. 2011. Antimicrobial resistance of Listeria monocytogenes isolates from food and the environment in France over a 10-year period. Applied and Environmental Microbiology 77, 2788-2790.

Haenni M., Saras E., Chaussière S., Treilles M. and Madec J.-Y. (2011). ermB-mediated erythromycin resistance in Streptococcus uberis from bovine mastitis in France. The Veterinary Journal, 189: 356-358.

Haenni M., Galofaro L., Ponsin C., Bes M., Laurent F. and Madec J.-Y. (2011) Staphylococcal bovine mastitis in France: enterotoxins, resistance and the human Geraldine methicillin-resistant Staphylococcus aureus clone. Journal of Antimicrobial Chemotherapy, 66 (1): 216-225

Haenni M, P. Châtre, S. Boisset, A. Carricajo, M. Bes, F. Laurent, JY. Madec. (2011). Staphylococcal nasal carriage in calves: multi-resistant S. sciuri and immune evasion cluster (IEC) genes in methicillin-resistant S. aureus ST398. Journal of Antimicrobial Chemotherapy, 66 (8): 1927-1928.

Le Devendec, L., Bouder, A., Dheilly, A., Hellard, G., Kempf, I., 2011, Persistence and Spread of qnr, Extended-Spectrum Beta-Lactamase, and ampC Resistance Genes in the Digestive Tract of Chickens. Microb Drug Resist 17, 129-134.

Le Hello, S., Hendriksen, R.S., Doublet, B., Fisher, I., Nielsen, E.M., Whichard, J.M., Bouchrif, B., Fashae, K., Granier, S.A., Jourdan-Da Silva, N., Cloeckaert, A., Threlfall, E.J., Angulo, F.J., Aarestrup, F.M., Wain, J., Weill, F.X. 2011. International spread of an epidemic population of Salmonella enterica serotype Kentucky ST198 resistant to ciprofloxacin. Journal of Infectious Diseases 204, 675-684.

Madec J.-Y., Doublet B., Ponsin C., Cloeckaert A. and Haenni M. (2011) Extended-spectrum beta-lactamase blaCTX-M-1 gene carried on an IncI1 plasmid in multidrug-resistant Salmonella enterica serovar Typhimurium in cattle in France. Journal of Antimicrobial Chemotherapy, 66 (4): 942-944.

Sakwinska O., Morisset D., Madec J.-Y., Waldvogel A., Moreillon P. and Haenni M.. (2011). Link between genotype and antimicrobial resistance in bovine-mastitis Staphylococcus aureus: comparison between Swiss and French isolates along the Rhône valley. Applied and Environmental Microbiology, 77: 3428-3432

31

Sanders P., Bousquet-Melou A., Chauvin

C., Toutain P.L., 2011. Utilisation des antibiotiques en élevage et enjeux de santé publique. INRA Productions Animales, 24, 199-204.

Veldman, K., L. M. Cavaco, D. Mevius, A. Battisti, A. Franco, N. Botteldoorn, M. Bruneau, A. Perrin-Guyomard, T. Cerny, C. de Frutos Escobar, B. Guerra, A. Schroeter, M. Gutierrez, K. Hopkins, A. L. Myllyniemi, M. Sunde, D. Wasyl, and F. M. Aarestrup. 2011. International collaborative study on the occurrence of plasmid-mediated quinolone resistance in Salmonella enterica and Escherichia coli isolated from animals, humans, food and the environment in 13 European countries. Journal of Antimicrobial Chemotherapy 66:1278-1286. Zeitouni, S., Kempf, I., 2011, Fitness Cost of Fluoroquinolone Resistance in Campylobacter coli and Campylobacter jejuni. Microb Drug Resist 17, 171-179.