intA PCR

In the early 2000s it was recognised that false positive test results were occurring in the gelatin gel test when it was applied in Merino flocks in the New England region of NSW in which there was no or doubtful clinical evidence of virulent footrot. This was leading to unnecessary quarantine and considerable hardship for some farmers. 

Cheetham et al. (2006) used Southern blot analysis on 595 isolates of D. nodosus from 124 farms on which sheep had benign or virulent footrot to test for the presence of the intA gene. They found that D. nodosus strains which were stable in the gelatin gel test and contained the intA gene could cause virulent footrot. They developed a polymerase chain reaction (PCR) to detect the intA gene. It could be used on  DNA extracted from cultured bacteria (Cheetham et al., 2006). The intA PCR test was subsequently evaluated independently using a wide range of D. nodosus samples from other geographic regions. The correlations observed in the New England were not confirmed (Dhungyel et al., 2013). Based on more than 2000 D. nodosus samples collected from 12 flocks of sheep with clinically virulent footrot, 64% of isolates were classified as virulent in the elastase test, compared to 91% in the gelatin gel test and 41% in the intA test; only about 21% of the isolates were virulent in all 3 tests. 

The intA DNA test has been applied in NSW as an adjunct to the gelatin gel test to distinguish benign footrot from virulent footrot in the face of positive (stable) results in the gelatin gel test.

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