RESEARCH Figure 2. Agarose gel electrophoresis of integrase gene poly- merase chain reaction (PCR) amplification products. A: PCR prod- ucts of class 1 integrase gene intI1. Lane M; molecular marker; lanes 1 and 19: no template (negative) control; lanes 2 and 20: positive control (In2); lanes 3–36: multiple drug–resistant isolates. B: PCR products of class 2 integrase gene intI2. Lane M: molecu- lar marker; lanes 1 and 19: no template control (negative) control; lanes 2 and 20: positive control (Tn7); lanes 3–36: multidrug- resistant isolates. itive (Figure 2B). This particular isolate (isolate 29) was obtained in the Presidio region and had a 2,600-bp variable region (Figure 3B). Nucleotide sequencing identified two distinct gene cassettes, namely, the sat-1 and aadA genes, which code for streptothricin acetyl transferase, and amino- glycosede adenyltransferase, respectively (Figure 5). Discussion Antimicrobial resistance in human pathogens has become a major public health issue. Resistant organisms have been isolated from a number of natural and man- made environments (6,19,20). In natural environments, resistant organisms can be indigenous or introduced through natural or anthropogenic causes (21,22). Integron gene sequences have been identified as a primary source of resistance genes and are suspected to serve as reservoirs of antimicrobial resistance genes within microbial popula- tions (1,2,23,24). Previous studies along the Texas-Mexico border have shown that fecal contamination of the Rio Grande does occur (7,25). The isolation of 322 E. coli iso- lates from irrigation water and associated sediments fur- ther confirms that fecal wastes are affecting this body of water. Previous studies have reported that municipal and animal wastes regularly harbor multidrug-resistant E. coli strains (6,26,27). In this study, 18% of the isolates were resistant to cephalothin. These results are similar to those from a recent survey of U.S. rivers, which found cefo- taxime (a third-generation, cephalosporin-resistant, gram- negative bacterium) to range from 16% to 96% across 22 rivers (19). The higher frequency of isolation of resistant strains from the El Paso region compared with the other, less urbanized sampling locations is not surprising since the effluent from a number of wastewater treatment plants enters the river at that region (W. McElroy, unpub. data; 28). Previous studies with sludge and septic tank wastes showed relatively high levels of antimicrobial resistance in E. coli (6). The precise sources of the E. coli isolates used in this study could not be identified because of technical limitations in source tracking (29). Class 1 and class 2 integron gene sequences were found within these E. coli isolates. Together, they accounted for 5 (16%) of 32 multidrug-resistant isolates characterized in this study. This prevalence was higher than that reported by Rosser et al. (30), who showed that 3.6% of gram-neg- ative bacteria in an estuarine environment contained the class 1 integron. Three of the four class 1 integron-bearing E. coli in this study contained the nucleotide sequence of the spectinomycin-streptomycin resistance gene aadA1 (31). Resistance to streptomycin was not observed in these isolates, but resistance to the closely related kanamycin was seen. These results are similar to those reported by Zhao et al. (3), who identified that the aadA gene trans- ferred to a strain of Hafnia alvei but did not report resist- ance to streptomycin or spectinomycin. These researchers attributed their findings to the inefficient expression of the inserted gene cassette by the integron promoter. Previous studies have also shown that the antimicrobial resistance phenotype can be modulated once these strains are exposed to specific environmental conditions (32). The aadA gene cassette is not novel in class 1 inte- grons. Earlier work by Zhao et al. (3) and Bass et al. (24) has shown that the aadA gene is highly conserved among Shiga toxin–producing and avian clinical E. coli isolates, respectively. The only class 2 integron-bearing strain iso- lated in this study also contained the aadA gene in addition to the sat-1 gene, which codes for resistance to kanamycin, a finding in agreement with the phenotypic expression. Figure 3. A: Polymerase chain reaction (PCR) amplification prod- ucts with primers targeted against the class 1–specific conserved sequences. Lane 1: no template control; lane 2: positive control (In2); lane 3: Escherichia coli isolate 16; lane 4: E . coli isolate 19; lane 5: E. coli isolate 21; lane 6: blank; lane 7: E. coli isolate I-6. B: PCR amplification products with primers targeted against the class 2–specific conserved sequences. Lane 1: no template con- trol; lane 2: positive control (In2); lane 3: E. coli isolate 29. 824 Emerging Infectious Diseases • Vol. 9, No. 7, July 2003