Transduction by bacteriophage P22 in nonsmooth mutants of Salmonella typhimurium

The general transducing phage P22 attacks only smooth (S) Salmonella with O antigen 12, determined by the oligosaccharide repeating unit constituting the distal part of the somatic lipopolysaccharide (LPS) side chain; non-S mutants, whose LPS contain few or no O repeating units, appear to be resistant. Auxotrophic non-S mutants of Salmonella typhimurium LT2 were tested as transductional recipients. Some transductants (0.5 to 5% as many as from S recipients) were obtained from most semirough recipients, either of class D (presumed leaky rouA mutants) or of a class due to mutation near his (presumed leaky rouB mutants), and from recipients lacking uridine diphosphogalactose epimerase or phosphomannose isomerase. Transductants were not obtained from several rouA, rouB, "heptose-negative," and glucose-1-transferase mutants, nor from most semirough class C mutants, whose LPS side chains each bear a single O oligosaccharide unit. Most transductants evoked from non-S recipients by temperate (c(+)) phage P22 were nonlysogenic, and virulent P22.c2 phage was about as effective as P22.c(+) in transduction to non-S recipients; probably all P22 transducing particles neither lysogenize nor kill. The extended-host-range mutant P22h gave qualitatively similar results,but evoked 5- to 30-fold more transductants from some non-S recipients than did P22. Probably, the LPS of non-S mutants susceptible to transduction contains a few O-specific oligosaccharide units, conferring a slight ability to adsorb P22 and a greater ability to adsorb P22h.     

Transport in bacteriophage P22-infected Salmonella typhimurium.

There was rapid efflux of L-leucine, L-phenylalanine, and alpha-methyl-D-glucoside after infection of Salmonella typhimurium with the clear plaque mutant C1 of phage P22. The efflux was similar to that observed with cyanide or arsenate treatment except that there was partial recovery in the case of phage infection and almost complete recovery under the condition of lysogeny. There was no efflux after infection with the temperature-sensitive mutant ts16C1 at nonpermissive temperature. Superinfection of superinfection exclusion negative lysogen (sie A minus sie B minus) with C1 led to efflux, whereas the efflux was much less on superinfection of sie A+ Sie B+ lysogen. These results indicate that an effective injection process is enough to cause depression in the cellular transport processes.     

Sequence of the genome of Salmonella bacteriophage P22

The sequence of the nonredundant region of the Salmonella enterica serovar Typhimurium temperate, serotype-converting bacteriophage P22 has been completed. The genome is 41,724 bp with an overall moles percent GC content of 47.1%. Numerous examples of potential integration host factor and C1-binding sites were identified in the sequence. In addition, five potential rho-independent terminators were discovered. Sixty-five genes were identified and annotated. While many of these had been described previously, we have added several new ones, including the genes involved in serotype conversion and late control. Two of the serotype conversion gene products show considerable sequence relatedness to GtrA and -B from Shigella phages SfII, SfV, and SfX. We have cloned the serotype-converting cassette (gtrABC) and demonstrated that it results in Salmonella serovar Typhimurium LT2 cells which express antigen O1. Many of the putative proteins show sequence relatedness to proteins from a great variety of other phages, supporting the hypothesis that this phage has evolved through the recombinational exchange of genetic information with other viruses.     

Patterns of transcription in bacteriophage P22-infected Salmonella typhimurium.

Two peaks of RNA synthesis (early and late) are directed by bacteriophage P22 in lytic infections of Salmonella typhimurium. Late RNA synthesis is not seen in P22 23- infections; neither early nor late RNA synthesis occurs in P22 24- infections. Genes 23 and 24 of P22 appear to be analogous to genes Q and N of lambda, respectively.     

Effect of spermidine on bacteriophage P22 infection.

The effect of spermidine on phage P22 infection of Salmonella typhimurium has been found to depend on the time of addition of spermidine with respect to the time of addition of the phage and also on the composition of the growth medium. If spermidine was added prior to or within a short time after infection, the cells survived. Under this condition the invading DNA appeared to remain trapped in the cell membrane, and there was no expression of the phage genome. If spermidine was added after the initiation of the infective process, the replication of the phage was inhibited but the cells did not survive. If spermidine was added after DNA synthesis was over, there was no effect of spermidine on phage multiplication. Spermidine was found to affect phage DNA synthesis but not host DNA synthesis.     

Mapping of rfa Genes in Salmonella typhimurium by ES18 and P22 Transduction and by Conjugation

Loci termed rfa, determining biosynthesis of somatic lipopolysaccharide core, have been mapped in Salmonella typhimurium LT2. The smooth-specific phage P22 co-transduced two leaky rfa alleles with cysE and with pyrE; one of the leaky alleles is perhaps rfaG, and the other is an unidentified gene concerned with synthesis of the heptose-containing part of the core. The lipopolysaccharide-indifferent phage ES18 (or its variant ES18.h1) co-transduced rfaF, rfaG, rfaL, rfa(R-res-1), and rfa(R-res-2) alleles with cysE and with pyrE, at rates indicating the order cysE–rfaF–(rfa[R-res-1], rfa[r-res-2], rfaL)–rfaG–pyrE. One proven (and two suspected) rfaE alleles and five proven rfaH alleles were not co-transduced with cysE or pyrE. Hfr crosses indicated that the proven rfaE allele mapped between serA and strA.     

Inhibition and eradication of Salmonella Typhimurium biofilm using P22 bacteriophage,EDTA and nisin

Abstract

P22 phage >10⁵ PFU ml^?1 could be used to inhibit Salmonella Typhimurium biofilm formation by 55–80%. Concentrations of EDTA >1.25?mM and concentrations of nisin >1,200?µg ml^?1 were also highly effective in reducing S. Typhimurium biofilm formation (≥96% and ≥95% reductions were observed, respectively). A synergistic effect was observed when EDTA and nisin were combined whereas P22 phage in combination with nisin had no synergistic impact on biofilm formation. Triple combination of P22 phage, EDTA and nisin could be also used to inhibit biofilm formation (≥93.2%) at a low phage titer (10² PFU ml^?1), and low EDTA (1.25?mM) and nisin (9.375?µg ml^?1) concentrations. A reduction of 70% in the mature biofilm was possible when 10⁷ PFU ml^?1 of P22 phage, 20?mM of EDTA and 150?μg ml^?1 of nisin were used in combination. This study revealed that it could be possible to reduce biofilm formation by S. Typhimurium by the use of P22 phage, EDTA and nisin, either alone or in combination. Although, removal of the mature biofilm was more difficult, the triple combination could be successfully used for mature biofilm of S. Typhimurium.     

Further physical characterization of deletion and substitution mutants affecting the control of lysogeny in bacteriophage P2

Summary A deletion of phage P2, del6 (L.E. Bertani, 1980), thought to remove the structural gene int, and a deletion/substitution, vir94, thought to remove genes int, C and cox, were mapped by electron microscopy, using the heteroduplex technique.Four independent deletion/substitution mutations, all affecting the regulatory region of P2, were compared in all possible combinations with the same technique: two showed sequence homology in their substitution DNA. The results confirm the model proposed for the origin of these mutants, analogous to that for the origin of transducing variants in phage , but suggest in first approximation that the exchange between the P2 DNA and the chromosome of the host bacterium may occur at several different bacterial sites.A map of the regulatory region of P2, based on all data available from the study of deletions and insertions, is presented.     

Growth of Salmonella bacteriophage P22 in Escherichia coli dna(Ts) mutants.

Salmonella bacteriophage P22 grows in two deoxyribonucleic acid initiation mutants of Escherichia coli under nonpermissive conditions, dnaA and dnaC. Functional products of genes dnaE, dnaZ, lig, dnaK, and dnaG are indispensable for deoxyribonucleic acid replication of P22. In 11 E. coli dnaB mutants belonging to all phenotypic groups, phage were produced at 42 degrees C.