Abstract

The Salmonella potsdam group B bacteriophage P3, which grows in the rough variant strain of S. potsdam (SP2-R) and E. coli KCS11, KW117, and C+/L with similar efficiencies, was restricted by SP2-R after a passage though E. coli. The entrance of restricted E. coli-grown phage DNA into nonpermissive SP2-R cells was confirmed by the finding that after adsorption of restricted 32P-labeled P3-C1·C+/L to SP2-R and removal of the adsorbed phage heads from cells, 80.3% of the labeled DNA activity sedimented with the cells. Thus restricted P3 DNA enters SP2-R cells and is probably cleaved by internal enzymes as in other systems.

With one-step growth experiments to investigate the P3 modification and restrictions, it was revealed that P3 grew more efficiently in SP2-R with burst sizes on SP2-R, E. coli, KCS11, KW1177, and C+/L of 182, 27, 24, and 91 respectively. After one growth cycle of P3·S in E. coli, the proportion of progeny unrestricted by SP2-R probably represented those having unrestricted parental, P3·S DNA that has been conserved.

The latent period, burst size, and proportion of P3 able to lyse SP2-R, after a growth cycle in E. coli KCS11 and KW1177 cultures grown overnight at 45 C, resembled those of phage grown in cells incubated at 37 C. The burst size of P3 in E.coli C+/L, grown at the elevated temperature, increased from 91 to 242, indicating the elevated temperature actually enhanced phage production in this strain. In contrast, the P3 burst size in SP2-R grown overnight at 45 C was about 20% of that in SP2-R grown at 37 FC. Phage P3 from a growth cycle in E. coli was similarly restricted by SP2-R indicator cultures grown overnight at 37 C and 45 C. This thermostability of the SP2-R restriction function was confirmed by the finding that SP2-R cells heated five minutes at 46-54 C still restricted phage P3 KW1177, and probably P3 KCS11 and P3 C+/L also. Starving and heating SP2-R in .0125 M CaCI2 for increasing times at elevated temperatures also failed to reduce the restriction.

SP2-R, E. coli KCS11, KW1177, and C+/L cultures ultraviolet-irradiated to < 3% survival still retained 35-90% of their phage producing capacity. SP2-R phage capacity was most resistant and revealed an S-shaped, multicomponent inactivation curve. Irradiation of recipient SP2-R cultures with up to 6,000 ergs/mm2 did not reduce the restriction of E. coli-grown P3·KCS11, P3·KW177, and P3·C+/L. Irradiation of E. coli prior to the P3·S growth cycle did not alter the proportion of unrestricted phage lysing SP2-R, but did reduce the burst size by one third to one-half and increase the latent periods, which indicated that the affected progeny phage DNA still carried the specificity or lack of specificity to be restricted.

Methionine deprivation of the auxotroph SP2-R met-, infected with P3-C1, did not affect the EOP of the phage on SP2-R compared to unrestrictive E. coli. UV-induction of methionine deprived SP2-3 met- cells also led to production of P3 phage which plated at similar EOP on SP2-R compared with the EOP on unrestrictive E. coli. These results did not disclose a role of methionine in the modification P3·S DNA. Methionine deprivation of E. coli KCS101 met- cells did not affect the restriction of P3·KCS101 in SP2-R or its final titers in the starved cells. A role of methylation in P3 specificity is, however, not precluded by these results.

LLU Discipline

Microbiology

Department

Microbiology

School

Graduate School

First Advisor

Leonard R. Bullas

Second Advisor

Robert L. Nutter

Third Advisor

R. Bruce Wilcox

Degree Name

Master of Science (MS)

Degree Level

M.S.

Year Degree Awarded

1971

Date (Title Page)

1-1971

Language

English

Library of Congress/MESH Subject Headings

Salmonella Phages; Escherichia coli.

Type

Thesis

Page Count

vii; 63

Digital Format

PDF

Digital Publisher

Loma Linda University Libraries

Usage Rights

This title appears here courtesy of the author, who has granted Loma Linda University a limited, non-exclusive right to make this publication available to the public. The author retains all other copyrights.

Collection

Loma Linda University Electronic Theses and Dissertations

Collection Website

http://scholarsrepository.llu.edu/etd/

Repository

Loma Linda University. Del E. Webb Memorial Library. University Archives

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