Isolation of Glycopeptide-Resistant Enterococci from Chickens in Japan

Avoparcin is a glycopeptide antibiotic that has been used as a growth-promoting agent for food animals in many countries, except the United States and Canada. In human medicine, glycopeptide-resistant enterococci (GRE) have become an increasingly serious problem in the treatment of nosocomial infections (6, 8, 10). In Europe, GRE have been isolated from animals, meat, the environment, and healthy humans outside hospitals (1, 2, 4, 7, 9), suggesting the possibility of transmission of GRE from food animals to humans via the food chain (2–4). In Japan, two glycopeptides have been approved as feed additives for chickens—avoparcin in 1985 and orienticin in 1994—but no papers have been published on the isolation of GRE from chickens in Japan. Then, a nationwide survey was conducted to isolate GRE from the fecal droppings of chickens on farms in Japan.

Prior to isolation of enterococci from fecal droppings, the distribution of avoparcin from the manufacturers to chicken farms was traced for 1 year across the country. Avoparcin was confirmed to have been brought into 24 prefectures in the period from December 1995 to November 1996. Fecal droppings were sampled from 39 broiler farms in these 24 prefectures. Fecal droppings of chickens not exposed to avoparcin were sampled from 11 broiler farms and 35 layer farms in 23 prefectures where no avoparcin had been distributed in the same period. Fecal samples were taken during a 1-week period from 11 December to 18 December 1996. Enterococci were isolated on bile esculin azide agar plates. MICs were determined for avoparcin, orienticin, and vancomycin by the agar dilution method with Mueller-Hinton agar.

Two hundred and sixty-three enterococci were isolated from the 35 broiler farms confirmed to have used avoparcin. Eight (3.0%) of them were GRE, originating from three (8.6%) farms. Of these eight strains, seven were identified as Enterococcus faecalis and the remaining one was identified as E. faecium. Five of the seven E. faecalis strains originated from the same farm, and the remaining two strains originated from another farm. All of the GRE were proved by PCR to harbor the vanA gene. No GRE were found among 295 enterococci isolated from broiler and layer farms not exposed to avoparcin (Table 1).

TABLE 1.

In vitro susceptibilities to avoparcin, orienticin, and vancomycin of enterococci isolated from fecal droppings of chickens

Sourcea (no. of isolates) and antibiotic	MIC (μg/ml)			No. of resistant isolates (%)
	Range	50%	90%
Broiler farms
Using avoparcin (263)
Avoparcin	0.78–>100	1.56	3.12	8 (3.0)
Orienticin	0.05–≥50	0.2	0.78	8 (3.0)
Vancomycin	0.39–>100	0.39	3.12	8 (3.0)
Not using avoparcin (81)
Avoparcin	0.78–3.12	1.56	1.56
Orienticin	0.05–1.56	0.2	0.78
Vancomycin	0.39–3.12	0.78	3.12
Layer farms not using avoparcin (214)
Avoparcin	0.78–3.12	1.56	1.56
Orienticin	0.05–1.56	0.2	0.78
Vancomycin	0.39–3.12	0.78	1.56

^aUse or lack of use of avoparcin is indicated for the period of December 1995 to November 1996. 

In Japan, the amounts of avoparcin recently marketed were 8,902 kg in 1993, 5,214 kg in 1994, 1,840 kg in 1995, and 986 kg in 1996, accounting for 2.78, 1.87, 0.68, and 0.40% of all growth-promoting antibiotics, respectively (5). Even though the consumption of avoparcin was smaller than that in Denmark (1) and has decreased each year, avoparcin use was associated with the presence of GRE in the feces of chickens after only a decade of use. The presence of GRE in the fecal droppings of chickens in this survey led to the ban of avoparcin and orienticin in food animals on 18 March 1997 in Japan.