Prevalence of tir gene subtypes in Escherichia coli isolates from pigs.

colony characteristics of isolates were noted, including haemolysis on blood agar and texture (smooth, rough, mucoid) on Tergitol-7 agar, for routine diagnostic procedures. Infections were considered to be potentially dual or multiple when colonies with different haemolytic patterns or morphology were observed on one plate. For each culture, all isolates with different phenotypes were selected for PCR analysis. PCR for tir gene subtypes was carried out as previously described by China and others (1999) and Nielsen and Andersen (2003) with the primers shown in Table 1. PCR for the Shiga toxin gene was also carried out as previously described by Kong and others (1999). The PCRs were performed in triplicate. Control DNA from reference strains was included in each reaction. The primer pair for each tir subtype yielded PCR products of the expected size for the control strains and field isolates. The results of the analysis of tir gene subtypes are shown in Table 2. None of the AEEC isolates studied carried the gene for Shiga toxin. The results of this study show the presence of various combinations of eae and tir subtype genes. The combination of eaeγ and tirγ is the most common in E coli isolated from human beings, while the pathotype eaeβ and tirβ, seems to be most frequently associated with diarrhoea in calves (China and others 1999). In these pigs, the eaeβ gene was associated with the tirθ gene in 14 of 59 isolates. Homologous combinations were also present: eaeβ with tirβ in two isolates and eaeγ with tirγ in three isolates. Four subtypes of tir were identified by PCR analysis of the AEEC isolates from diarrhoeic Korean pigs. The prevalence of tir subtypes in E coli from the preweaned pigs was similar to that from the postweaned pigs. Fourteen of the isolates could not be typed because they did not react with the PCR primers used in this study. These results suggest that some eae-positive E coli may possess another unique subtype or may have different sequences in their subtypes that prevent their recognition by PCR. All the AEEC strains used in this study were isolated from the ileum, because all the samples were submitted for the Prevalence of tir gene subtypes in Escherichia coli isolates from pigs