Description and validation of a novel technique to study the bacterial flora of the teat duct of ewes

We describe a new technique that may be used for studying the bacterial flora of the teat duct in live ewes, and we present evidence regarding its accuracy, safety and repeatability. A sterile, plastic, 20 G catheter is used for sampling. The catheter stylet is taken out and the plastic catheter cut with a sterile blade to a length of 2 mm. In order to ensure accurate and consistent cutting of the catheter at the desired length, a sterilized ruler is placed beside the catheter. The whole procedure is carried out under aseptic conditions, on sterilized surgical screens. Before sampling, a thorough disinfection with povidone iodine scrub solution of the teat apex and the lower (1 cm) part of the teat skin is carried out. The catheter is held by the investigator from the cannula hub; then, it is inserted into the teat, rolled around the internal teat wall, in order to sample the mucosa and then withdrawn. Material collected on the tip of the catheter is plated onto bacteriological media. The sensitivity of the technique compared to a method that detects the bacterial flora of the teat after slaughter of the animals (scratching of the teat mucosa by means of a scalpel blade) was 99.1% for field samples and 100% for experimental samples; specificity of the technique was 90.7 and 100%, respectively, kappa coefficient between results of the two methods was 0.9045. The safety of the technique was assessed in an investigation, where ewes were repeatedly (eight times) sampled by means of this technique, throughout their lactation; in no case was clinical or subclinical mastitis recorded. Finally, the repeatability of the method was assessed in 37 paired samples, obtained 30-40 min apart; a 100% match was recorded between bacteriological results of the two samplings of each pair. We conclude that the method can be useful in research studies, in order to obtain information regarding bacterial ecology and dynamics inside the teat of ewes.