Multidrug-resistant (MDR) Pseudomonas aeruginosa infections pose a serious health threat. Bacteriophage–antibiotic combination therapy is a promising candidate for combating these infections. A 5-phage P. aeruginosa cocktail, PAM2H, was tested in combination with antibiotics (ceftazidime, ciprofloxacin, gentamicin, meropenem) to determine if PAM2H enhances antibiotic activity. Combination treatment in vitro resulted in a significant increase in susceptibility of MDR strains to antibiotics. Treatment with ceftazidime (CAZ), meropenem, gentamicin, or ciprofloxacin in the presence of the phage increased the number of P. aeruginosa strains susceptible to these antibiotics by 63%, 56%, 31%, and 81%, respectively. Additionally, in a mouse dorsal wound model, seven of eight mice treated with a combination of CAZ and PAM2H for three days had no detectable bacteria remaining in their wounds on day 4, while all mice treated with CAZ or PAM2H alone had ~107 colony forming units (CFU) remaining in their wounds. P. aeruginosa recovered from mouse wounds post-treatment showed decreased virulence in a wax worm model, and DNA sequencing indicated that the combination treatment prevented mutations in genes encoding known phage receptors. Treatment with PAM2H in combination with antibiotics resulted in the re-sensitization of P. aeruginosa to antibiotics in vitro and a synergistic reduction in bacterial burden in vivo.

Synthetic peptides encoding protective pathogen-derived epitopes represent -in principle -an ideal approach to T-cell vaccination. Empirically, however, these strategies have not been successful. In the current study, we profiled the early activation of CD8 1 T cells by MHC class I-restricted peptide immunization to better understand the biology of this response. We found that CD8 1 T cells proliferated robustly in response to low doses of short synthetic peptides in PBS, but failed to acquire effector function or form memory populations in the absence of the TLR ligand CpG. CpG was unique among TLR ligands in its ability to enhance the response to peptide and its adjuvant effects had strict temporal requirements. Interestingly, CpG treatment modulated T-cell expression of the surface receptors PD-1 and CD25, providing insight into its possible adjuvant mechanism. The effects of CpG on peptide immunization were dramatically enhanced in the absence of B cells, demonstrating a unique system of regulation of T-cell responses by these lymphocytes. The results reported here provide insight into the complex response to a simple vaccination regimen, as well as a framework for a rational peptide-based vaccine design to both exploit and overcome targeted aspects of the immune response.

Liver-stage antigen 1 (LSA1) is expressed byPlasmodium falciparum liver-stage antigen 1 (LSA1) is expressed exclusively in the liver stage of parasite development (17). The native LSA1 protein has a large central repeat region containing over 83 slightly degenerate 17-amino-acid repeat segments, flanked by two highly conserved N-and C-terminal regions. The lsa1 gene encodes a 230-kDa protein whose expression begins shortly after sporozoite invasion of the liver hepatocyte and increases rapidly with liver-stage development (13, 17). The LSA1 protein has been described as forming a flocculent mass surrounding the thousands of developing parasite nuclei and pseudocytomeres and is microscopically observed localized between the plasmalemma and the parasitophorous vacuole membranes inside the infected hepatocyte cytoplasm. Later in development the LSA1 protein has been detected adhering to emerging mature liver merozoites (13).It has been demonstrated that complete resistance to malaria infection can be induced in rodents and humans by prior injection, by needle or mosquito bite, of gamma-irradiated sporozoites (␥-spz) and that the mechanism of immunity is, in large part, due to major histocompatibility complex class I-restricted CD8 ϩ gamma interferon (IFN-␥)-secreting T cells directed against antigens expressed by malaria-infected hepatocytes (8-10, 12, 16, 17, 22, 23). Furthermore, the effector cells of this immune response target primarily liver-stage developing merozoites and not sporozoites before their entry into hepatocytes. In mice, the developing attenuated liver-stage parasites resulting from the irradiated sporozoites must be present at the time of normal sporozoite challenge for immunity to be manifested (16,22,27,39). In humans, the protection induced by the gamma-irradiated sporozoite model is effective against homologous and heterologous parasite strain challenge, indicating that the antigen(s) responsible is relatively conserved. The role of specific anti-LSA1 immune responses in this protection is evident. It has been reported that human volunteers exposed to a protective dose of P. falciparum irradiated sporozoites develop LSA1 peptide-specific proliferative T cells (17,20). In addition, epidemiologic studies have reinforced the association of LSA1 to disease resistance or severity among individuals living in areas where malaria is endemic. Studies of antibody (Ab) and CD4 ϩ or CD8 ϩ T-cell responses measured by antigenic specific cellular proliferation, cytokine production (primarily IFN-␥ and interleukin-10 [IL-10]), and presence of cytotoxic lymphocytes has been demonstrated in residents of various areas where malaria is endemic (6,11,25,35). Although these data should be interpreted cautiously given the differing ethnicities and ages of the subjects and variety of immunologic assays used, it appears that IFN-␥ and IL-10

A potential concern with bacteriophage (phage) therapeutics is a host-versus-phage response in which the immune system may neutralize or destroy phage particles and thus impair therapeutic efficacy, or a strong inflammatory response to repeated phage exposure might endanger the patient. Current literature is discrepant with regard to the nature and magnitude of innate and adaptive immune response to phages. The purpose of this work was to study the potential effects of Staphylococcus aureus phage K on the activation of human monocyte-derived dendritic cells. Since phage K acquired from ATCC was isolated around 90 years ago, we first tested its activity against a panel of 36 diverse S. aureus clinical isolates from military patients and found that it was lytic against 30/36 (83%) of strains. Human monocyte-derived dendritic cells were used to test for an in vitro phage-specific inflammatory response. Repeated experiments demonstrated that phage K had little impact on the expression of pro- and anti-inflammatory cytokines, or on MHC-I/II and CD80/CD86 protein expression. Given that dendritic cells are potent antigen-presenting cells and messengers between the innate and the adaptive immune systems, our results suggest that phage K does not independently affect cellular immunity or has a very limited impact on it.