A simplified method has been devised for typing Pseudomonas aeruginosa by pyocin production. Pyocins are produced as strains grow overnight in Trypticase soy broth (without glucose) plus 1% potassium nitrate. Because P. aeruginosa can use nitrate instead of oxygen as a terminal electron acceptor, mechanical shaking is not necessary, nor is induction by mitomycin C. Pyocins can now be produced in screw-cap tubes in a water bath or incubator. A total of 250 strains were tested as possible pyocin indicators, which included 60 strains already used in pyocin-typing systems. The final set contained 18 indicators which were chosen because (i) they had clear positive or clear negative reactions, thus eliminating reactions difficult to read, (ii) they had few zones due to bacteriophage lysis, and (iii) they were most sensitive in differentiating clinical isolates of P. aeruginosa. The final typing method was tested in several studies and the results were clear; thus definitive epidemiological conclusions could be made. Because it is simple to perform and easily automated, the new method should have application in many hospitals; however, it should be used only in carefully planned epidemiological studies. The method and its application are described in detail, and some pitfalls are discussed. Nosocomial infections caused by Pseudomonas aeruginosa continue to be a problem in many hospitals (3, 5, 8, 12, 16, 23). Within any institution there can be hundreds of different strains because it is often found in the human gut, and because it can survive and multiply in moist environments of the hospital. In any epidemiological study, a sensitive technique is required to compare these numerous strains. A number of methods have been used to differentiate P. aeruginosa in epidemiological studies. These epidemiological markers include

, nine cases of Pseudomonas aeruginosa septicemia occurred in a high-risk nursery. The epidemiology of the outbreak was studied by pyocin production, pyocin sensitivity, serological typing, antibiotic susceptibility, and phenotypic properties such as colonial morphology, pigment, and hemolysis. Ten isolates of P. aeruginosa were recovered from 9 newborn infants and from 13 environmental sources. Twenty-one of the 23 isolates had identical pyocin production patterns against 60 different indicator strains and were of the same serotype. These 21 isolates were designated as the "outbreak strain"; the other 2 isolates had no epidemiological significance. The results of pyocin sensitivity, antibiotic susceptiblity tests, and phenotypic properties were dissimilar. They would yield incorrect epidemiological conclusions if used alone. The outbreak strain dissociated in vitro and these phenotypic changes accounted for the variable results by the latter three typing methods. Although the precise mode of introduction of the organism into the nursery could not be determined in retrospect, the epidemiological data strongly suggested that one infant contracted a P. aeruginosa infection, and this strain spread throughout the nursery by means of contaminated resuscitation equipment.

In April 1971, nine cases of Pseudomonas aeruginosa septicemia occurred in a high-risk nursery. The epidemiology of the outbreak was studied by pyocin production, pyocin sensitivity, serological typing, antibiotic susceptibility, and phenotypic properties such as colonial morphology, pigment, and hemolysis. Ten isolates of P. aeruginosa were recovered from 9 newborn infants and from 13 environmental sources. Twenty-one of the 23 isolates had identical pyocin production patterns against 60 different indicator strains and were of the same serotype. These 21 isolates were designated as the “outbreak strain”; the other 2 isolates had no epidemiological significance. The results of pyocin sensitivity, antibiotic susceptibility tests, and phenotypic properties were dissimilar. They would yield incorrect epidemiological conclusions if used alone. The outbreak strain dissociated in vitro and these phenotypic changes accounted for the variable results by the latter three typing methods. Although the precise mode of introduction of the organism into the nursery could not be determined in retrospect, the epidemiological data strongly suggested that one infant contracted a P. aeruginosa infection, and this strain spread throughout the nursery by means of contaminated resuscitation equipment.

Good yields of pyocin are obtained when Pseudomonas aeruginosa is grown in Trypticase soy broth (without glucose) (BBL) plus 1% potassium nitrate. As a result, pyocin production for routine epidemiological typing can be done in screw-cap tubes, without mechanical agitation or mitomycin C induction.

No abstract