Detection of streptomycin resistance in Erwinia amylovora strains isolated from apple orchards in Chihuahua, Mexico

Abstract

Fire blight, one of the most severe diseases of apple and pear, is caused by the bacterium Erwinia amylovora. One control method is the use of antibiotics like streptomycin; however, streptomycin is the only antibiotic registered to control fire blight. A total of 107 E. amylovora strains were isolated from apple orchards located in Cuauhtemoc and Guerrero, Chihuahua, two major apple-producing areas in Mexico, showing 40 and 24 % streptomycin-resistant strains, respectively. The identification of E. amylovora strains was performed by polymerase chain reaction (PCR) amplification of a 900-bp region located within the non-transferable pEA29 plasmid and by amplification of a specific 1,269-bp region located on the E. amylovora chromosome. The 107 isolates tested carried the pEA29 plasmid, and 36 % of the isolates from both locations showed high resistance to streptomycin at levels that ranged from 200 to ≥1,000 μg ml−1 streptomycin. The strA-strB and aadA genes, which encode enzymes that inactivate streptomycin, and a mutation in codon 43 of the rpsL gene that confers high resistance to the antibiotic were examined to determine the mechanism of streptomycin resistance. In total, 95 % of the resistant strains showed a single base pair mutation in codon 43 of the rpsL gene, causing an amino acid substitution in ribosomal protein S12. The presence of strA-strB and aadA genes or the rpsL mutation was not identified in the other 5 % of resistant strains, suggesting the existence of a new streptomycin resistance mechanism in E. amylovora.