The present study was undertaken to define the standard conditions for growth of bacteriophage 7V on its host Pseudomonas aeruginosa strain PS-7 and to determine the factors which affect the quantity and quality of plaques in the plaque count assay. Observations from the single-step growth experiment and single-burst experiment are also included. Plaque count assays were performed under various environmental conditions. Conditions were selected as “standard” if they yielded: 1) relative maximum number of infective centers per ml of stock 7V phage, 2) clear, haloed plaques at least 2.0 mm in diameter, and 3) reproducible assays limited only by the sampling error. These conditions are: 1. fresh NBYE or NBYE agar for the growth medium 2. NYBE or buffered salts solution for diluents 3. Physiologically young cells in the log phase between 1-5 X 10⁸ bacteria/ml 4. Stock and diluted stock suspensions stored at refrigerator temperatures. Adsorption rate experiments which measured both unadsorbed phage and infective centers were performed in minimal media, minimal media supplemented with organic and and ionic cofactors, and complete media. Although overnight lysates of PS-7 in minimal media produced a high titer of phage, the rate of adsorption of phage 7V in PS-7 was extremely slow in minimal media. Addition of tryptophan caused a decrease in free phage without a corresponding increase in infective centers. Casamino acids plus tryptophan caused an increase in the velocity of the adsorption reaction which was less than the rate of adsorption of phage 7V to its host PS-7 in NBYE. In NBYE 90 percent of the initial phage were adsorbed in 5 minutes, but the recovery of phage as free phase of infective centers was not equal to the input of phage. These results suggest that this system requires a cofactor, organic, ionic, or both, in order that adsorption of phage 7V to its host PS-7 proceed at a maximum rate. And it further suggests that the incidence of abortive infection in this system is high. In this particular system under standard conditions it appears that the size of the plaques is controlled mainly by environmental factors, while the relative number of plaques is a characteristic of the system.
| Identifer | oai:union.ndltd.org:pdx.edu/oai:pdxscholar.library.pdx.edu:open_access_etds-1770 |
| Date | 01 May 1969 |
| Creators | Schnider, Shirley Phillips |
| Publisher | PDXScholar |
| Source Sets | Portland State University |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Dissertations and Theses |
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