Genetic diversity among VRE isolates from Swedish broilers with the coincidental finding of transferrable decreased susceptibility to narasin.

AIMS: In this study, the molecular diversity among clones of vancomycin resistant Enterococcus faecium with vanA gene (VRE) is investigated. The aims were to better understand why one clone is predominant in Swedish broiler production and to better assess the potential for zoonotic gene transfer from the different clones. METHODS AND RESULTS: Twenty-six isolates were separated into 11 clones. Vancomycin resistance was transferrable from the predominant and five minority clones. Decreased susceptibility to narasin was co-transferred with vancomycin resistance in four clones, including the predominant. The plasmid addiction system axe-txe was not detected, and the ω-ε-ζ system was detected in one of the minority clones but was not co-transferred with vancomycin resistance. CONCLUSIONS: The results do not explain why one clone is predominant among VRE in Swedish broiler production but confirms the potential for zoonotic spread of vancomycin resistance genes. The near absence of investigated plasmid addiction systems indicates that they do not play an important role in the epidemiology of VRE in Swedish broiler production. The finding that decreased susceptibility to narasin can be co-transferred with the vanA gene indicates that the use of narasin might play a role in the persistence of vancomycin resistance in enterococci colonizing Swedish broilers. SIGNIFICANCE AND IMPACT OF THE STUDY: This is, to our knowledge, the first report of transferrable decreased susceptibility to narasin.

Inter-batch contamination and potential sources of vancomycin-resistant Enterococcus faecium on broiler farms.

1. Vancomycin-resistant Enterococcus faecium (VRE(fm)) has recently spread among Swedish broiler farms. The objectives were to investigate VRE(fm) persistence within barns between flocks, and to determine whether day-old chicks, feed or forklift trucks used for loading crates could be identified as a means of transmission. 2. Faeces were collected for selective culture from 12 farms (9 culture-positive, 3 culture-negative as determined by prior monitoring), and samples were collected from the barn environment before and after cleaning and disinfection, from forklift tyres, hatcheries and feed. 3. VRE(fm) was isolated only from previously known VRE(fm)-positive farms. The proportions of culture-positive environmental samples were 75% (9 out of 9 farms) prior to and 31% (7 out of 9 farms) after cleaning/disinfection. Five out of 6 samples from forklift tyres were culture-positive. No VRE(fm) were isolated from feed or hatcheries. The majority of 27 vanA gene positive isolates showed similar banding patterns by SmaI restriction digestion and pulse-field gel electrophoresis. No consistent differences were observed regarding management between VRE(fm)-positive and negative farms. 4. We conclude that VRE(fm) contaminates barns and remains present between flocks. Forklift trucks are one possible means of transmission between farms.

Reflection paper on MRSA in food-producing and companion animals: epidemiology and control options for human and animal health.

The scope of this reflection paper was to review the latest research on the risk of MRSA infection and colonization in animals. Attention focused on occurrence, risk factors for colonization and infection, and human contact hazard for livestock, horses, and companion animals. Whereas the clonal relationship between MRSA strains of CC398 is straightforward in livestock this is less obvious in horses. Small companion animals typically share MRSA strains that seem to exchange with a human reservoir. Management and therapeutic options have been suggested for livestock, horses, companion animals, as well as instructions on safety measures for persons in contact with animals. Conclusions were drawn with emphasis on future research activities, especially to confirm the apparent evolution of the organism and to demonstrate efficiency of control strategies.

Spread without known selective pressure of a vancomycin-resistant clone of Enterococcus faecium among broilers.

OBJECTIVES: The aim of this paper was to describe an increased occurrence of vancomycin-resistant enterococci (VRE) in Swedish broilers since 2000 and to investigate the genetic relatedness of isolates. METHODS: Caecal content from slaughtered broilers was cultured for VRE on medium supplemented with vancomycin (16 mg/L). Species identification, antibiotic susceptibility determination, vancomycin resistance genotyping, multilocus sequence typing (MLST) and characterization of Tn1546 were performed. RESULTS: The proportion of VRE-positive samples increased gradually from <1% in 2000 to slightly over 40% in 2005. Between 2005 and 2006, the proportion of VRE-positive samples decreased and between 2006 and 2007, it was stable at just below 30%. All isolates tested were Enterococcus faecium and carried the vanA gene. A majority of the isolates had similar antibiograms, the same MLST sequence type and Tn1546 transposon. CONCLUSIONS: The proportion of VRE-positive samples from broilers has increased since 2000, and this is due to the spread of one major clone. Moreover, this has taken place in an environment without any obvious selective pressure.

Antimicrobial resistance in Escherichia coli isolated from bitches with pyometra and from urine samples from other dogs.

To assess whether the rates of antimicrobial susceptibility in bacteria isolated from the urine of dogs with urinary tract infections are similar to those of bacteria isolated from bitches with pyometra, the antimicrobial resistance of Escherichia coli isolated from the two groups were determined and compared. The samples were collected in Sweden between April 2002 and March 2003, and potential changes over time were assessed by comparing the results with corresponding data from 1991 to 1993. Among 80 isolates of E coli from cases of pyometra, the proportions that were resistant to the antimicrobials used in canine practice were generally low (ampicillin 10 per cent, enrofloxacin 4 per cent, gentamicin 0 per cent, streptomycin 5 per cent, sulfamethoxazole 8 per cent, tetracycline 4 per cent and trimethoprim 2 per cent) and similar to the proportions reported previously. Significantly lower proportions of resistance were recorded among the pyometra isolates than among 92 isolates from urine samples submitted by animal hospitals to ampicillin (P=0.04), streptomycin (P=0.002) and tetracycline (P=0.03), but there were no differences between the pyometra isolates and 113 isolates from urine samples submitted by animal clinics.

Antimicrobial susceptibility of equine isolates of Actinobacillus spp. and identification of beta-lactamases in some strains.

A total number of 149 Actinobacillus strains isolated from clinical samples (73 strains) and from the oral cavity of healthy horses (76 strains) were tested for their susceptibility to 17 antimicrobial substances. The antibiograms were generally very similar between the various strains and no differences could be clearly correlated to either phenotype or source of isolates. However, when tested against penicillin, ampicillin, trimethoprim-sulfa, and streptomycin, small groups of strains with what appeared to be acquired resistance could be identified. Eight of the penicillin-resistant strains were found to produce beta-lactamase. The beta-lactamases appeared to be bound tightly to the cell wall, thereby frustrating further characterization by isoelectric focusing. Plasmids of approximately 3 kb were found in four out of seven beta-lactamase-producing strains submitted to plasmid analysis.

Antimicrobial susceptibility of Swedish, Norwegian and Danish isolates of Clostridium perfringens from poultry, and distribution of tetracycline resistance genes.

This study was undertaken to determine the in vitro susceptibility of Clostridium perfringens, isolated from poultry to antimicrobials used in poultry production. The minimal inhibitory concentration (MIC) of eight antimicrobials, including the ionophoric coccidiostat narasin, was determined for 102 C. perfringens isolates, 58 from Sweden, 24 from Norway and 20 from Denmark. Susceptibility to each antimicrobial compound was determined by broth microdilution. The isolates were obtained from broilers (89), laying hens (9) and turkeys (4), affected by necrotic enteritis (NE) or by C. perfringens associated hepatitis (CPH), and from healthy broilers. All strains, regardless of origin, proved inherently susceptible to ampicillin, narasin, avilamycin, erythromycin and vancomycin. A low frequency of resistance to virginiamycin and bacitracin was also found. Resistance to tetracycline was found in strains isolated in all three countries; Sweden (76%), Denmark (10%) and Norway (29%). In 80% of the tetracycline-resistant isolates, the two resistance genes tetA(P) and tetB(P) were amplified by PCR whereas in 20% only the tetA(P) gene was detected. No tetM gene amplicon was obtained from any of the tetracycline-resistant isolates. The uniform susceptibility to narasin revealed in this study shows that the substance can still be used to control clostridiosis. In this study, C. perfringens also showed a low degree of resistance to most other antimicrobials tested. Despite the small amounts of tetracycline used in poultry, a considerable degree of resistance to tetracycline was found in C. perfringens isolates from Swedish broilers.

The usage of veterinary antibacterial drugs for mastitis in cattle in Norway and Sweden during 1990-1997.

The prescribing patterns and annual incidence of use of antibacterial drugs for the treatment of mastitis in cattle in Norway and Sweden during the period 1990-1997 were estimated from drug wholesaler statistics. Although the drugs included in this study are also used in other species and/or other indications, mastitis in cattle is by far the most-common indication for their use. We used these sales figures to evaluate trends in the use of antibacterial drugs and changes in the incidence of treatment in bovine mastitis in Norway and Sweden. To facilitate comparisons (correcting for differences in activity and dosage) between the relative proportions of antibacterial drugs used in bovine mastitis, we introduced defined daily dose cow (DDDcow) as unit of measurement. Tentative DDDcow for the various injectable drugs were derived from doses recommended in Norway and Sweden. For intramammary drugs administered in the form of single-dose applicators, one applicator was defined as the DDDcow. The prescribing patterns of antibacterial drugs in bovine mastitis in Norway and Sweden during the study period seemed to be influenced by treatment policies, substances and formulations approved and treatment cost; length of the withdrawal period also seemed to play a role.

Antimicrobial resistance in staphylococci from canine pyoderma: a prospective study of first-time and recurrent cases in Sweden.

In a prospective study involving eight veterinary clinics during 1995 and 1996, samples from first-time and recurrent cases of canine pyoderma were collected by a needle technique. Three hundred and ninety-four staphylococci were isolated and their susceptibility to various antimicrobial drugs was assessed by a microdilution technique. Resistance to macrolides, lincosamides, fusidic add, tetracycline and streptomycin was significantly more common in isolates from the recurrent cases than from the first-time cases; 20 per cent of the isolates from the first-time cases were resistant to three or more of the antimicrobials tested, compared with 45 per cent of those from the recurrent cases. Coresistance between macrolide-lincosamides, tetracyclines and streptomycin was common. No resistance to penicillinase-stable beta-lactams was observed. A comparison with earlier studies indicated that there had been a marked increase in resistance during the previous five years.

Antibacterial drugs prescribed for dogs and cats in Sweden and Norway 1990-1998.

The usage of veterinary antibacterial drugs in dogs and cats in Sweden and Norway for the period 1990-1998 was investigated by use of drug wholesalers' statistics. Additionally, usage of human antibacterial drugs in these species in Sweden was investigated by use of prescription data for the period 1996-1998. On average, more than 50% of the prescribed veterinary antibacterials in Sweden were beta-lactam antibiotics. In Norway, about 75% of the preparations prescribed for dogs and cats contained sulfonamides and trimethoprim. Furthermore, the prescription data from Sweden showed a reduced usage of human antibacterials prescribed for dogs and cats since the beginning of the 1980s. Approximately 20% of the prescribed packages for dogs in the years 1996-1998 were human approved drugs. The corresponding figure for cats was 13%. The differences between the countries in the choice of antibacterial drugs can be explained by differences in the availability of approved preparations during the study period. The consumption of veterinary antibacterials in dogs and cats in Sweden during the period was in the range of 3% to 8% of the total use of veterinary antibacterials. The corresponding figures in Norway were in the range of 3% to 7%. It is of vital importance to study usage patterns of antibacterial drugs in dogs and cats in surveillance and control of bacterial resistance, but also in discussions of therapeutic appropriateness. Therefore, further research is needed in this area.

Assessment of phagocytic capacity and opsonic activity in blood and mammary secretions during lactation in sows.

The objective of this study was to determine whether phagocytic capacity and opsonic activity in blood and mammary secretions of sows are impaired at parturition compared with later on during lactation. The study comprised eight primiparous sows (Landrace x Yorkshire) free from clinical signs of disease. Blood and mammary secretion samples were collected within 48 h of parturition and 7 and 16 days after parturition. Numbers and proportion of polymorphonuclear neutrophils (PMN) were determined in blood and mammary secretions. Phagocytic capacity was assessed in whole blood and in a cell suspension derived from mammary secretions. Opsonic activity was assessed in serum and i cell-depleted, skimmed mammary secretions. The two assays were based on chemiluminescence, both having zymosan and Escherichia coli as target particles. Numbers and proportion of PMN in mammary secretions were higher (P < 0.05) at parturition than later on during lactation. A parturition, phagocytic capacity in cell suspensions derived from mammary secretions was higher for both (P < 0.05) and E. coli (P < 0.1). However, when phagocytic capacity was related to the number of PMN in the suspension no such difference was observed. The opsonic activity in cell-depleted, skimmed mammary secretions at parturition was lower (P < 0.05) for zymosan but not for E. coli. None of the described variations were reflected in blood or serum. The findings of this study do not unequivocally support the theory that an immune suppression at parturition in the sow can help explain the increased incidence of coliform mastitis at that time.

Actinomyces canis sp. nov., isolated from dogs.

Three strains of a previously undescribed catalase-positive Actinomyces-like bacterium were isolated from dogs. Biochemical testing and PAGE analysis of whole-cell proteins indicated that the strains were phenotypically highly related to each other but different from previously described Actinomyces and Arcanobacterium species. Sequencing of 16S rRNA showed that the unknown bacterium represents a new subline within a cluster of species which includes Actinomyces hyovaginalis, Actinomyces georgiae, Actinomyces meyeri, Actinomyces odontolyticus, Actinomyces radingae and Actinomyces turicensis. On the basis of phenotypic evidence and 16S rRNA sequence divergence levels (greater than 5% with recognized Actinomyces species) it is proposed that the unknown strains from canine sources be classified as a new species with the name Actinomyces canis sp. nov. The type strain of Actinomyces canis is CCUG 41706T (= CIP 106351T).

Actinomyces bowdenii sp. nov., isolated from canine and feline clinical specimens.

Four strains of a previously undescribed Actinomyces-like bacterium were isolated from canine and feline clinical specimens. Phenotypic studies indicated the strains were members of the genus Actinomyces, and most closely resembled Actinomyces viscosus serotype I and Actinomyces slackii. Comparative 16S rRNA gene sequencing studies demonstrated the unknown bacterium constitutes a new subline within a group of Actinomyces species, which includes Actinomyces bovis, the type species of the genus. Based on phylogenetic and phenotypic evidence it is proposed that the unknown bacterium be classified as Actinomyces bowdenii sp. nov. The type strain of Actinomyces bowdenii is CCUG 37421T.

Efficacy of trimethoprim-sulfadoxine against Escherichia coli in a tissue cage model in calves.

Tissue cages implanted subcutaneously in calves were infected with Escherichia coli. Twenty-four hours later, the calves were treated either with single doses of 2.5 + 12.5 or 5 + 25 mg/kg trimethoprim (TMP) + sulfadoxine (SDX) or with five doses of 7.5 + 37.5 mg/kg TMP + SDX at 12-h intervals. In addition, one cage in each of three calves in the highest dose group was infected 3 h after initiation of treatment. Untreated calves were kept as controls. Concentrations of TMP and SDX in plasma and tissue cage fluid (TCF) and counts of viable bacteria in TCF were determined. In the highest dose group, concentrations of TMP in TCF remained above the minimum inhibitory concentration of the test strain for 94-101 h and peak to minimum inhibitory concentration (MIC) ratio was close to 10. In spite of this, an effect of treatment was noted only in cages infected after initiation of treatment. In vitro studies and analysis of thymidine content in serum and TCF from calves suggest that levels of thymidine in TCF are high enough to antagonize the antibacterial effect of TMP. The results indicate that soft tissue infections in secluded infection sites of calves are refractory to treatment with TMP + SDX.