Bacterial Virus Lambda Gpd-Fusions to Cathelicidins, α- and β-Defensins, and Disease-Specific Epitopes Evaluated for Antimicrobial Toxicity and Ability to Support Phage Display.

We showed that antimicrobial polypeptides, when translated as gene fusions to the bacteriophage lambda capsid decoration protein gpD, formed highly toxic molecules within , suggesting that they can retain their antimicrobial activity conformation when fused to gpD. These include gpD-fusions to human and porcine cathelicidins LL37 and PR39, β-defensins HBD3 and DEFB126-Δ (deleted for its many COOH-terminal glycosylation sites), and α-defensin HD5. Antimicrobial toxicity was only observed when the peptides were displayed from the COOH-terminal, and not the NH-terminal end, of gpD.

Complementation Studies of Bacteriophage λ O Amber Mutants by Allelic Forms of O Expressed from Plasmid, and O-P Interaction Phenotypes.

λ genes O and P are required for replication initiation from the bacteriophage λ origin site, λ, located within gene . Questions have persisted for years about whether O-defects can indeed be complemented . We show the effect of original null mutations in O and the influence of four origin mutations (three are in-frame deletions and one is a point mutation) on complementation.

The Bacteriophage Lambda CII Phenotypes for Complementation, Cellular Toxicity and Replication Inhibition Are Suppressed in cII-oop Constructs Expressing the Small RNA OOP.

The temperate bacteriophage lambda (λ) CII protein is a positive regulator of transcription from promoter , a component of the lysogenic response. The expression of was examined in vectors devoid of phage transcription-modulating elements. Their removal enabled evaluating if the expression of the small RNA OOP, on its own, could suppress CII activities, including complementing for a lysogenic response, cell toxicity and causing rapid cellular loss of ColE1 plasmids.

Lambda display phage as a mucosal vaccine delivery vehicle for peptide antigens.

Bacteriophage are structurally stable in the gastro-intestinal tract and have favorable traits of safety, stability, ease of production, and immunogenicity. These attributes make them potential candidates as oral vaccine delivery vehicles but little is known about their capacity to induce mucosal immune responses in the small intestine. Whole body imaging of mice confirmed lambda bacteriophage (LP) were distributed throughout the gastro-intestinal tract 24 h after oral delivery.

Lambda gpP-DnaB Helicase Sequestration and gpP-RpoB Associated Effects: On Screens for Auxotrophs, Selection for Rif(R), Toxicity, Mutagenicity, Plasmid Curing.

The bacteriophage lambda replication initiation protein P exhibits a toxic effect on its Escherichia coli (E. coli) host, likely due to the formation of a dead-end P-DnaB complex, sequestering the replicative DnaB helicase from further activity. Intracellular expression of P triggers SOS-independent cellular filamentation and rapidly cures resident ColE1 plasmids.

Phage Lambda P protein: trans-activation, inhibition phenotypes and their suppression.

The initiation of bacteriophage λ replication depends upon interactions between the oriλ DNA site, phage proteins O and P, and E. coli host replication proteins. P exhibits a high affinity for DnaB, the major replicative helicase for unwinding double stranded DNA.

A CI-independent form of replicative inhibition: turn off of early replication of bacteriophage lambda.

Several earlier studies have described an unusual exclusion phenotype exhibited by cells with plasmids carrying a portion of the replication region of phage lambda. Cells exhibiting this inhibition phenotype (IP) prevent the plating of homo-immune and hybrid hetero-immune lambdoid phages. We have attempted to define aspects of IP, and show that it is directed to repλ phages.

Dual expression system for assembling phage lambda display particle (LDP) vaccine to porcine Circovirus 2 (PCV2).

The bacteriophage lambda small capsid protein D forms trimers on the phage head. D-fusion polypeptides can be expressed from plasmids in E. coli and remain soluble without aggregation.

Immunogenicity of bacteriophage lambda particles displaying porcine Circovirus 2 (PCV2) capsid protein epitopes.

Phage lambda particles displaying four immunodominant regions of porcine Circovirus 2 (PCV2) capsid protein (LDP-D-CAP) was shown to be immunogenic in pigs. The immunodominant regions were fused to the carboxyl-terminal of lambda head protein D. Expression of D-CAP on lambda display particles was demonstrated by ELISA and Western blots.

NinR- and red-mediated phage-prophage marker rescue recombination in Escherichia coli: recovery of a nonhomologous immlambda DNA segment by infecting lambdaimm434 phages.

We examined the requirement of lambda recombination functions for marker rescue of cryptic prophage genes within the Escherichia coli chromosome. We infected lysogenic host cells with lambdaimm434 phages and selected for recombinant immlambda phages that had exchanged the imm434 region of the infecting phage for the heterologous 2.6-kb immlambda region from the prophage.

Polarity within pM and pE promoted phage lambda cI-rexA-rexB transcription and its suppression.

The cI-rexA-rexB operon of bacteriophage lambda confers 2 phenotypes, Imm and Rex, to lysogenic cells. Immunity to homoimmune infecting lambda phage depends upon the CI repressor. Rex exclusion of T4rII mutants requires RexA and RexB proteins.

Over-expression of rexA nullifies T4rII exclusion in Escherichia coli K(lambda) lysogens.

Dosage and relative cellular levels of RexA and RexB proteins encoded by the rexA-rexB genes of a lambda prophage are important for the Rex+ phenotype, which was nullified when greater RexA or RexB was provided than was necessary for the complementation of a rexA- or a rexB- prophage.

Blocking the T4 lysis inhibition phenotype.

Nonlysogenic Escherichia coli K cells exhibit a delay in lysis when infected by T4rII phage termed lysis inhibition (LIN). E. coli K cells expressing lambda rexB from either a prophage defective for rexA, or a multicopy plasmid supported T4rII infection, but prevented the establishment of LIN.

Bacteriophage lambda repressor allelic modulation of the Rex exclusion phenotype.

The sensitivity of delta red-gam delta ren mutants of bacteriophage lambda to Rex exclusion by lambda rexA+ rexB+ lysogens is modulated by the prophage cI repressor allele. We show the following: (i) lambda spi156 delta nin5 forms plaques on a cI+-rexA+-rexB+ lysogen with 10(5)-fold higher efficiency than on cI[Ts]-rexA+-rexB+ derivatives. (ii) The cI[Ts]857 allele augmentation of Rex exclusion is recessive to cI+.

Rex-centric mutualism.

We asked whether Rex exclusion encoded by a lambda prophage confers a protective or a cell-killing phenotype. We found that the Rex system can channel lysogenic cells into an arrested growth phase that gives an overall protective ability to the host despite some associated killing.