Revised pyocin typing method for Pseudomonas aeruginosa

Abstract

In the Gillies and Govan method of pyocin typing for Pseudomonas aeruginosa a cross-streaking technique was used, and 105 main types and 25 subtypes were identified by the patterns of inhibition observed on 13 indicator strains. Disadvantages of the technique included the need to remove test strain growth before application of the indicator strains, the 48-h period needed to obtain a result, and the inability to reliably type mucoid P. aeruginosa. Recent studies have enabled us to overcome these disadvantages and significantly improve the speed and application of pyocin typing. Our revised technique utilizes the same 13 indicator strains which are already used internationally. Test strains were rapidly applied to the surface of agar plates with a multiple inoculator. After incubation for 6 h and exposure to chloroform, the indicator strains were applied in agar overlays without prior removal of the test strain growth. After 18 h of incubation, the pyocin type was recognized by inhibition of particular indicator strains. Additionally, the activity of particulate (R and F) and nonparticulate (S) pyocins could be distinguished on the basis of inhibition zone size, which thus allowed further discrimination. The revised technique allows typing within 24 h, increases the number of identifiable types, and can be used to type mucoid strains.

Development of resistance in Pseudomonas aeruginosa obtained from patients with cystic fibrosis at different times

“…The P. aeruginosa strains were analyzed for different pyocin types by the spotting method as described by Fyfe et al. [4] and pulse field gel electrophoresis (PFGE) patterns by contour‐clamped homogeneous electric field electrophoresis as described earlier [5]. Genomic DNA was subjected to the rare‐cutting restriction enzyme SpeI .…”

Epidemiologic characterization of Pseudomonas aeruginosa in patients with cystic fibrosis

“…Pyocin typing was done by the spotting method as described by Fyfe et al [21]. This method is based on the inhibition of 13 P. aeruginosa indicator strains by pyocin produced by the patients' strains to be typed.…”

Engineering bacteriocin-mediated resistance against plant pathogenic bacteria in plants

“…All Soft agar overlay susceptibility assays. Soft agar overlay spot assays were performed using the method of Fyfe et al (43). Fifty microliters of test strain culture at mid-log was inoculated in 0.8 % soft agar and poured over a KB agar plate, as appropriate.…”