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Background In Denmark in 1994 and 1995 there was strong media attention focused on the use of antimicrobials in food animal production. Two aspects were emphasized. One was the perceived overuse of antibiotics in pig herds, in particular overuse of tetracycline. The other was the finding that use of the antimicrobial growth promoter avoparcin was associated with the occurrence of vancomycinresistant Enterococcus faecium in pigs and broilers (Bates et al. 1994; Klare et al. 1995; Aarestrup, 1995; Bager et al., 1997) and in foods of animal origin (Wegener et al., 1997).

Animals and humans constitute overlapping reservoirs of resistance determinants (Levy, 1997); and the concern was that indiscriminate use of antimicrobials in food animal production would, by way of food, result in transfer of resistance factors to humans (Corpet, 1988; Sorensen et al., 2001) where it might compromise treatment of infectious diseases. The concerns resulted in a number of government initiatives, among them measures to reduce veterinarian prescription of antimicrobials, measures against use of certain antimicrobial growth promoters and the establishment of the DANMAP monitoring and research programme.

The EU has procedures for centralised approval of feed additives, and only antimicrobials specifically approved for growth promotion may be used for this purpose. Until 1999, with the exception of tylosin, bacitracin and spiramycin, only antimicrobials not used in human therapy were registered for growth promotion. In May 1995, the Danish government banned the use of avoparcin, a move followed by an EU-wide ban in 1996. In 1998, a further Danish ban on the use of virginiamycin for growth promotion was instituted because of cross-resistance to Synercid, a new antibiotic marketed for the treatment of infections from vancomycin resistant E. faecium. This prompted a re-consideration by the EU Scientific Steering Committee of the public health aspects of the use for growth promotion of antimicrobials closely related to compounds used in human therapies. This resulted in a recommendation (Scientific Steering Committee, 1999) that no antimicrobial belonging to groups of compounds used in the treatment of animals or humans should be used for growth promotant purposes. This was in line with the recommendations given in the Swann report in 1969 (Anon., 1969). Thus, since mid-1999 the only antimicrobial growth promoters approved within the EU are the ionophores salinomycin and monensin, and the antibiotics flavomycin and avilamycin.

The January 1998 government ban on use of virginiamycin and the media debate that followed prompted the Danish food animal industries to discontinue all use of antimicrobials for growth promotion. This response to customer concerns became effective in broilers, pigs over 35 kg live weight and in cattle in February 1998 and in weaner pigs in the autumn of 1999. At present, antimicrobial growth promoters are not used in Denmark, even though the four agents mentioned above remain licensed within the EU. In 1997, a total of 106 tonnes of antimicrobials were used for growth promotion, compared with 57 tonnes of antimicrobials for treatment of disease in food animals.

The DANMAP programme was established to monitor trends in antimicrobial resistance in bacteria from food animals, foods and humans and to track usage of antimicrobials in animals and humans. This has made it possible to monitor the effects of these interventions on resistance. An important part of DANMAP is scientific research. All bacterial isolates included in the programme are stored in a strain collection with all identifiers. Combined with information about antimicrobial usage, this has provided a basis for important insights about the molecular epidemiology of antimicrobial resistance in food animals. Additional scientific studies (as yet not fully completed) were launched to document the effects of the withdrawal of antimicrobial growth promoters on animal morbidity and on the usage of antimicrobials for treatment of disease in animals.

The measures aimed at curbing veterinary use of antimicrobials included restrictions on the antibiotic quantities that could be prescribed or sold to a farmer at any one time, measures to limit the practitioner’s profits on drug sales, and the introduction of legislation making it illegal to use antibiotics for prophylaxis, except to treat a specific infection known to be present in the herd or flock. In other words the measures aimed at reducing the use of antimicrobials as cheap alternatives to good animal management.

The present paper describes the design of the part of DANMAP that monitors food animals and the effect of the discontinued use of antimicrobial growth promoters on resistance and on mortality, with main emphasis on broiler production.


Design of resistance monitoring

SAMPLING PROCEDURES

Food animals

Samples are collected from the normal population of slaughter animals as well as from the diseased population. Meat inspection staff collect samples of caecal contents (pigs), cloacal swabs (broilers) or rectal contents (young bulls) from slaughter animals. The number of samples per plant was established in proportion to the annual throughput of the plant. Only one sample per herd or flock is collected on any one sampling occasion. The pig and broiler slaughter plants included in the programme account for between 95 and 98% of the total national production of these species, while for cattle the figure is about 80%. Formal randomisation is not carried out, and while the procedure resembles a stratified sampling plan there is a bias toward inclusion of large herds in preference to small. The herd identity is recorded at sampling and when the data are processed, it is possible to ensure that any one herd is included only once per year.

Isolates of animal pathogens are collected as a systematic random sample among isolates from diagnostic submissions to the Danish Veterinary Laboratory or to diagnostic laboratories operated by the animal industries. Between them, these laboratories process close to 100% of diagnostic samples from food animals in Denmark. The bacteria included from food animals and the target number of isolates are shown in Table 1.



Table 1. Bacteria from food animals included in the DANMAP programme. Figures represent the target number of isolates.

All bacteria included in DANMAP are stored in a strain collection and are available for studies of, for example, the genetic basis for resistance phenotypes or for retrospective studies to determine the occurrence of resistance to new types of antimicrobials. A full set of epidemiological descriptors is available for each isolate (farm and animal species of origin and time and place of sample collection). From 2001, the susceptibility data for each isolate may be linked with data on antimicrobial usage on the farm of origin.


Susceptibility testing

Most susceptibility testing is carried out using broth microdilution in Sensititre (Trek Diagnostic Systems, Ltd). However, Campylobacter are tested using agar plate dilution. The antimicrobial panels used include representatives of the main groups of therapeutic agents as well as antimicrobial growth promoters. A complete list of the antimicrobials included in the panel may be found in the DANMAP reports (DANMAP, 2001).


MONITORING USAGE OF ANTIMICROBIALS

All antibiotics for treatment of animals are prescription-only medicines and obtainable exclusively through pharmacies. Veterinarians must obtain medicines for use in practice from a pharmacy. They can legally sell the medicines to farmers, however, their profit is by law limited to 5%, which is deemed sufficient to cover expenses. Consequently, about 80% of all medicines used in veterinary practice are sold as prescriptions, which are redeemed by farmers at a pharmacy.

Until 2000, recording use of antibiotics for treatment of animals was based on compulsory reporting to the Danish Medicines Agency (DMA) by importers and manufacturers of the quantities sold to wholesalers and pharmacies. Reporting from the top of the supply pyramid meant that information on usage in the individual animal species was not available.

In 2000 the VETSTAT programme was implemented to provide continuous usage data for all prescription medicine in food animals at the individual herd level. VETSTAT is based on electronic registration of relevant information from all prescriptions processed at pharmacies and transfer of the information to VETSTAT via the DMA. In addition, veterinarians report sales of medicines to the central database (Figure 1). Among the information recorded by VETSTAT is the identity of the farm receiving the medicine, the species of animal and the age group for which the medicine is intended as well as the reason for prescribing. The data are collated to a national monthly statistic on drug usage in animals. The statistic is published on the internet. The results are used in detailed studies of the association between antimicrobial consumption and occurrence of resistance, as well as analysis of prescribing patterns. However, the data will also be used to form a basis for advice to veterinarians and farmers about prudent use of antimicrobials.


Trends in resistance

Using the DANMAP monitoring programme, we have been able to demonstrate the effects of the avoparcin ban on occurrence of vancomycinresistant E. faecium (VRE) in broilers and pigs. In broilers, we saw a rapid decline as early as 1996 (one year after the ban) while in pigs there was no apparent effect (Bager et al., 1999). In pigs, however, a decline started in 1998, following the voluntary discontinuation of growth promoter use in pigs above 35 kg live weight. Molecular studies found that a likely explanation for the initial lack of effect was that vancomycin-resistant E. faecium from pigs were co-selected by the widespread use of tylosin for growth promotion (Aarestrup, 2000). Stopping use of other growth promoting antibiotics has also resulted in decline in resistance to these agents (Aarestrup et al., 2001). Studies have shown that changes in occurrence of VRE in broilers at slaughter are reflected in their occurrence on broiler carcasses (Figure 2). We have found that this association also applies to resistance to other antimicrobial growth promoters formerly used.

It is noteworthy, however, that while this reflects a reduced exposure of consumers to these types of resistance factors and a subsequent decreased rateof shedding VRE in humans (Klare et al., 1999),
studies using sensitive methods have demonstrated that VRE remain in farm environments and may be found on broiler carcasses 3-4 years after the avoparcin ban (Heuer et al., 2002; Borgen et al., 2001).





Figure 1. Flow of data and information in VETSTAT.





Figure 2. Probability of vancomycin resistance among Enterococcus faecium in broiler meat, predicted by occurrence in broilers at slaughter.



In broiler production the discontinuation of antimicrobial growth promoters did not affect mortality rate and productivity also remained the same. However, the feed conversion ratio increased marginally by 16 g of feed per kg bird produced (Emborg et al., 2001). This increased feed consumption was offset by savings on growth promoting antimicrobials. There have been no indications of increased usage of antimicrobials for treatment of disease in broilers and therefore no increase in morbidity.

The withdrawal in 1998 of growth promoters for pigs over 35 kg live weight resulted in a small, transient drop in daily weight gain, but had no effect on the feed conversion ratio. Overall, the withdrawal of growth promoters had no effect on productivity in this age group and there was no effect on animal morbidity. However, in the age group 7 to 35 kg there was both a decline in daily weight gain and increased mortality. This may also be transient, as recent data show declining mortality and increasing weight gain. The disease problems experienced by some herds have contributed to an increased usage of prescription medicines for treatment. The problems most often consist of post-weaning diarrhoea and diarrhoea caused by infection with Lawsonia intracellularis. In addition, Postweaning Multisystemic Wasting Syndrome (PMWS) is present in Danish pig herds and may have contributed to the problems observed. Overall, the usage of antimicrobials for therapy has gone up (see below), however, it is rather uncertain to what extent this can be explained by the withdrawal of antimicrobial growth promoters.

Figure 3 shows the trend in resistance among Escherichia coli from diagnostic submissions from broilers and pigs. For pigs, the isolates belong to serotype O149, most of them isolates from cases of diarrhoea in weaner pigs. Weaner pigs represent the group where most of the antimicrobials in pig herds are used, however there has been little change in the rate of resistance and little reflection of the rate of the increased usage of antimicrobials. It does, however, serve to illustrate the complex dynamics of the spread of resistant bacteria and demonstrates that factors other than selective pressure from antimicrobials are involved. For example, among E. coli from broilers, a similar proportion to that found in pigs is resistant to tetracycline, although only insignificant amounts of tetracycline are used in commercial broiler flocks.






Figure 3. Trend in resistance among Escherichia coli from diagnostic submissions from broilers and pigs. The broiler isolates are serotypes O2 and O78 and the pig isolates are serotype O149. Data: DANMAP 2000.



Trend in usage of antimicrobials

Phenotypic expressions of antimicrobial resistance may occur as a result of co-selection by unrelated compounds because of linked resistance genes or it may be due to cross-resistance to related antimicrobial agents. Therefore, resistance monitoring alone does not provide the information required for well-targeted intervention. Focused intervention to control antimicrobial resistance must be based on detailed information about the use of antimicrobials, in addition to resistance data. Figure 4 shows the trend in usage of antimicrobial agents in Denmark. There was a peak in usage of antimicrobials for animal therapy in 1994. The subsequent decline to a low of about 48 tonnes in 1996, down from a total of about 90 tonnes two years previously, is the result of the government interventions described above. During this 2-year period there were no general changes in health status in the food animal population and little change in the size of the population. It is therefore likely that the difference – just over 40 tonnes of active compound – represents overuse.

Since 1996, antibiotic usage in animals has increased steadily. At least in part, the increase may be explained by increased animal production, however, there may still be some overuse. The reason, of course, is that to a very large extent the decision about when to medicate is made by the farmer and is very much a subjective one. From 2000 to 2001 the increase was bigger than in previous years. In 2001 we changed the reporting system and there may be systematic differences. For example, the data for 2001 include all medicines prescribed for companion animals, including nonveterinary medicines.

Irrespective of this undesirable increase in antibiotic usage in pigs, the total quantity used in 2001 was about 73 tonnes, or the equivalent of 3.3 g per pig slaughtered. Recent figures published by the Department for Environment, Food, and Rural Affairs (Anon., 2002) indicate that in the UK usage in pigs is at least as high as 15 g per pig produced, and possibly higher.

In contrast to the situation in animals, consumption of antimicrobials in human medicine in Denmark has remained stable and is among the lowest in Europe (Figure 4).



Conclusion

Denmark has an active policy to control antimicrobial usage in humans as well as in food animals. In the latter, initiatives have included government bans on use of the growth promoters avoparcin and virginiamycin, bans that monitoring has shown to have been effective in reducing the exposure of consumers to bacteria carrying resistance to these two agents. Another measure has been controls on use of antimicrobials for therapy. These resulted in the reduction of usage from a total of just under 90 tonnes active compound in 1994 to 48 tonnes in 1996, most likely an elimination of overuse. In the second half of the 1990s usage has increased again, an increase greater than the increase in animal production.

From late 1999, the Danish food animal industries voluntarily stopped using antimicrobial growth promoters. While in the broiler industry this was implemented without any loss of productivity or increase in mortality, the discontinued use in weaner pigs has resulted in problems with post-weaning diarrhoea, contributing to an increased usage of antimicrobials for therapy. This increased usage has not yet led to a significant increase in antimicrobial resistance among E. coli from diagnostic submissions from weaner pigs.





Figure 4. Trends in usage of antimicrobials.



Acknowledgements

The DANMAP programme is jointly funded by the Ministry of Food, Agriculture and Fisheries and the Ministry of Health. The programme is a collaboration between the Danish Veterinary Laboratory, the Danish food Directorate, Statens Serum Institut and the Danish Medicines Agency. VETSTAT is funded solely by the Ministry of Food, Agriculture and Fisheries.



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