Abstract
Four streptomycin-resistant isolates of Xanthomonas oryzae pv. oryzae (YNA7-1, YNA10-2, YNA11-2, and YNA12-2) were examined via PCR amplification for the presence of class 1, class 2, and class 3 integrons and aadA1 and aadA2 genes, which confer resistance to streptomycin and spectinomycin. The class 1 integrase gene intI1 and the aminoglycoside adenylyltransferase gene aadA1 were identified in all four resistant isolates but not in 25 sensitive isolates. PCR amplifications showed that 7790-bp, 7162-bp, 7790-bp, and 7240-bp resistance integrons with transposition gene modules (tni module) in 3′ conserved segments existed in YNA7-1, YNA10-2, YNA11-2, and YNA12-2, respectively. Subsequent analysis of sequences indicated that the integrons of YNA7-1 and YNA11-2 carried three gene cassettes in the order |aacA3|arr3|aadA1|. The integron of YNA10-2 carried only |arr3|aadA1| gene cassettes. The integron of YNA12-2 lacked a 550-bp sequence including part of intI1 but it still carried |aacA3|arr3|aadA1| gene cassettes. The analysis of inactive mutants and complementation tests confirmed that the aacA3 gene conferred resistance to tobramycin, kanamycin, gentamicin and netilmicin; the arr3 gene conferred resistance to rifampicin; and the aadA1 gene conferred resistance to streptomycin and spectinomycin. The resistance phenotypes of the four isolates corresponded with their resistance gene cassettes, except that YNA7-1 and YNA12-2 did not show rifampicin resistance. Sequence comparison revealed that no gene cassette array in GenBank was in the same order as in the integrons of the four resistant isolates in this study and the aadA1, which was identical in the four resistant isolates, showed 99% identity with aadA1 sequences in GenBank. The result of a stability test showed that the resistance phenotype, the aadA1 gene, and the intI1 gene were completely stable in YNA7-1 and YNA12-2 but unstable in YNA10-2 and YNA11-2. To our knowledge, this is the first report of resistance integron in a phytopathogenic bacteria.
Highlights
Bacterial blight of rice, caused by Xanthomonas oryzae pv. oryzae, is a serious disease in many rice-growing regions of the world, including the south of China [1,2,3]
The test results showed that four isolates (0.75% of the total) of X. oryzae pv. oryzae from the same county in Yunnan Province were highly resistant to streptomycin and that the resistance mechanism could not be attributed to the occurrence of strA-strB genes or to the rpsL gene mutation previously determined to cause streptomycin resistance in phytopathogenic bacteria [11,12,13,14,15,16]
In agreement with the negative results shown for the three sensitive isolates in Table 2, none of the 22 additional sensitive isolates showed any amplification products, which indicated that the aadA1 gene in the class 1 integron might contribute to the streptomycin resistance in the four resistant isolates
Summary
Bacterial blight of rice, caused by Xanthomonas oryzae pv. oryzae, is a serious disease in many rice-growing regions of the world, including the south of China [1,2,3]. Bacterial blight of rice, caused by Xanthomonas oryzae pv. Oryzae, is a serious disease in many rice-growing regions of the world, including the south of China [1,2,3]. Bacterial blight causes at least 10% yield loss on susceptible rice varieties when the weather is conducive [3,4]. Bismerthiazol is the most commonly used bactericide for control of bacterial blight of rice in China [5], but bismerthiazol rapidly selects for bismerthiazol-resistant strains of X. oryzae pv. Streptomycin, an aminoglycoside antibiotic that has been widely used in treatment of bacterial diseases of humans and animals [8], is used to control bacterial blight of rice in China [9]. Oryzae were collected in the south of China to determine their susceptibility to streptomycin. The test results showed that four isolates (0.75% of the total) of X. oryzae pv. oryzae from the same county in Yunnan Province were highly resistant to streptomycin and that the resistance mechanism could not be attributed to the occurrence of strA-strB genes or to the rpsL gene mutation previously determined to cause streptomycin resistance in phytopathogenic bacteria [11,12,13,14,15,16]
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