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There is a broad interest in adapting live vaccine strains (LVS) for use as recombinant vaccines that can deliver heterologous antigens. The Salmonella enterica serovar Typhimurium SL1344 ΔwecA LVS contains a mutation in wecA that abrogates production of Enterobacterial common antigen. This ΔwecA strain is attenuated in vivo, persistently colonizes the host, and protects against both wild type and cross-Salmonella serovar lethal challenge in a murine model of salmonellosis. Given these characteristics, we hypothesized that the SL1344 ΔwecA strain could be used as a carrier for heterologous antigen expression. To test this hypothesis, SL1344 ΔwecA was engineered to express the Pseudomonas aeruginosa O11 O-antigen gene cluster. Intraperitoneal (IP) but not oral immunization of BALB/c mice with the heterologous expression strain protected against lethal P. aeruginosa intranasal (IN) challenge. Furthermore, IP immunization resulted in P. aeruginosa O11-specific Ig and IgG antibody production. Functional analysis of sera collected from the IP immunized mice showed antibody-mediated agglutination and opsonophagocytic activity against P. aeruginosa. En masse, these results indicate that the S. Typhimurium SL1344 ΔwecA strain expressing the P. aeruginosa O11 O-antigen gene cluster is able to induce a humoral immune response and to protect against lethal P. aeruginosa challenge. As such, the S. Typhimurium SL1344 ΔwecA LVS can likely serve as a vehicle for expression of a wide variety of heterologous antigens as a means to create recombinant vaccines.

Accidental ingestion of a foreign body is a rare occurrence in the unimpaired adult. The usual patient is aware of the ingestion event and seeks medical attention if needed. We report a case of accidental, unrecognized, fatal ingestion of a foreign body discovered during a postmortem examination. In situ examination revealed a small perforation of the terminal ileum by a perforating metal wire. Fibrinopurulent peritonitis was the immediate cause of death. The foreign body was later found to closely resemble the wire bristles of a barbeque grill brush that family members brought to a family/clinician/pathologist conference. There are few previous case reports of ingestion of a grill brush wire, all of which were nonfatal. Health care workers, including pathologists, should be aware of this potential ingestible foreign body given the widespread use of barbeque grills in food preparation. Additionally, manufacturers and users of grill brushes should be well aware of this ingestion hazard.

The Gram-negative pathogen Helicobacter pylori is increasingly more resistant to the three major antibiotics (metronidazole, clarithromycin and amoxicillin) that are most commonly used to treat infection. As a result, there is an increased rate of treatment failure; this translates into an overall higher cost of treatment due to the need for increased length of treatment and/or the requirement for combination or sequential therapy. Given the rise in antibiotic resistance, the complicated treatment regime, and issues related to patient compliance that stem from the duration and complexity of treatment, there is clearly a pressing need for the development of novel therapeutic strategies to combat H. pylori infection. As such, researchers are actively investigating the utility of antimicrobial peptides, small molecule inhibitors and naturopathic therapies. Herein we review and discuss each of these novel approaches as a means to target this important gastric pathogen.

Helicobacter pylori has developed antimicrobial resistance to virtually all current antibiotics. Thus, there is a pressing need to develop new anti- H. pylori therapies. We recently described a novel oligo-acyl-lysyl (OAK) antimicrobial peptidomimetic, C 12 K-2β 12 , that shows potent in vitro bactericidal activity against H. pylori . Herein, we define the mechanism of action and evaluate the in vivo efficacy of C 12 K-2β 12 against H. pylori after experimental infection of Mongolian gerbils. We demonstrate using a 1- N -phenylnaphthylamine (fluorescent probe) uptake assay and electron microscopy that C 12 K-2β 12 rapidly permeabilizes the bacterial membrane and creates pores that cause bacterial cell lysis. Furthermore, using nucleic acid binding assays, Western blots, and confocal microscopy, we show that C 12 K-2β 12 can cross the bacterial membranes into the cytoplasm and tightly bind to bacterial DNA, RNA, and proteins, a property that may result in inhibition of enzymatic activities and macromolecule synthesis. To define the in vivo efficacy of C 12 K-2β 12 , H. pylori -infected gerbils were orogastrically treated with increasing doses and concentrations of C 12 K-2β 12 1 day or 1 week postinfection. The efficacy of C 12 K-2β 12 was strongest in animals that received the largest number of doses at the highest concentration, indicating dose-dependent activity of the peptide ( P < 0.001 by analysis of variance [ANOVA]) regardless of the timing of the treatment with C 12 K-2β 12 . Overall, our results demonstrate a dual mode of action of C 12 K-2β 12 against the H. pylori membrane and cytoplasmic components. Moreover, and consistent with the previously reported in vitro efficacy, C 12 K-2β 12 shows significant in vivo efficacy against H. pylori when used as monotherapy. Therefore, OAK peptides may be a valuable resource for therapeutic treatment of H. pylori infection.

CD62L (L-selectin) is a key regulator of T cell trafficking, and its surface expression on activated T cells is modulated to control T cell access to lymph nodes after acute infections. In memory T cells, CD62L is the most frequently used marker to define central memory T cells, a population that provides enhanced protection against most, but not all, pathogens. Early access of CD62Lpos effector T cells to lymph nodes has been proposed to result in preferential central memory T cell differentiation, but direct proof for the involvement of lymph node homing in memory T cell differentiation is lacking. In this study, we show that central memory lineage commitment in CD8 T cells is unaltered by enhanced entry into lymph nodes as a result of constitutive CD62L expression, and that equal numbers of effector and central memory CD8 T cells develop in the absence of CD62L-mediated lymph node trafficking. Our results suggest that CD62L is not a deterministic marker of central memory T cell differentiation, thus providing new insight into the process of memory CD8 T cell generation.

Normal adrenocortical activity is necessary for electrolyte regulation and the maintenance of cardiovascular function. Although chronic adrenal insufficiency generally presents with the gradual onset of a set of characteristic symptoms and signs, the more sudden loss of adrenal activity can present with acute, rapidly progressive cardiovascular dysfunction that can be fatal if not recognized and treated promptly. We herein describe a patient who had most of his adrenal tissue removed during resection of metastatic renal carcinoma, conventional clear cell type, with much of the remaining adrenal tissue undergoing necrosis during or shortly after surgery. Although the patient appeared to be stable and progressing adequately well, he died suddenly 2 days postoperatively. When the gross autopsy findings suggested the possibility of adrenal insufficiency, clinical laboratory assessment of adrenocortical activity was sought. Analysis of stored antemortem serum samples and of blood obtained at autopsy demonstrated a progressive decrease in cortisol levels which, in this stressed postsurgical patient, proved fatal. The use of both antemortem and postmortem blood in the demonstration of acute adrenal insufficiency at autopsy is discussed.

Immunity induced by the 19-kDa fragment ofPlasmodium yoeliimerozoite surface protein 1 (MSP119) is dependent on high titers of specific antibodies present at the time of challenge and a continuing active immune response postinfection. However, the specificity of the active immune response postinfection has not been defined. In particular, it is not known whether anti-MSP119antibodies that arise following infection alone are sufficient for protection. We developed systems to investigate whether an MSP119-specific antibody response alone both prechallenge and postchallenge is sufficient for protection. We were able to exclude antibodies with other specificities, as well as any contribution of MSP119-specific CD4+T cells acting independent of antibody, and we concluded that an immune response focused solely on MSP119-specific antibodies is sufficient for protection. The data imply that the ability of natural infection to boost an MSP119-specific antibody response should greatly improve vaccine efficacy.

It is thought that both helper and effector functions of CD4(+) T cells contribute to protective immunity to blood stage malaria infection. However, malaria infection does not induce long-term immunity and its mechanisms are not defined. In this study, we show that protective parasite-specific CD4(+) T cells were depleted after infection with both lethal and nonlethal species of rodent PLASMODIUM: It is further shown that the depletion is confined to parasite-specific T cells because (a) ovalbumin (OVA)-specific CD4(+) T cells are not depleted after either malaria infection or direct OVA antigen challenge, and (b) the depletion of parasite-specific T cells during infection does not kill bystander OVA-specific T cells. A significant consequence of the depletion of malaria parasite-specific CD4(+) T cells is impaired immunity, demonstrated in mice that were less able to control parasitemia after depletion of transferred parasite-specific T cells. Using tumor necrosis factor (TNF)-RI knockout- and Fas-deficient mice, we demonstrate that the depletion of parasite-specific CD4(+) T cells is not via TNF or Fas pathways. However, in vivo administration of anti-interferon (IFN)-gamma antibody blocks depletion, suggesting that IFN-gamma is involved in the process. Taken together, these data suggest that long-term immunity to malaria infection may be affected by an IFN-gamma-mediated depletion of parasite-specific CD4(+) T cells during infection. This study provides further insight into the nature of immunity to malaria and may have a significant impact on approaches taken to develop a malaria vaccine.

Acid-sensing ion channel type 1 (ASIC1), which may play contribute to synaptic plasticity, memory, fear conditioning and ischemic brain injury, is found primarily in brain; but it is also expressed...

The adhesion molecule L-selectin is cleaved rapidly from the surface of activated leukocytes by tumor necrosis factor-alpha converting enzyme, a cell surface metalloprotease, and also undergoes slower constitutive shedding in unactivated cells. The structural features that render it susceptible to shedding are poorly understood. We therefore analyzed the shedding of a series of mutant and chimeric L-selectin molecules. Although murine L-selectin is cleaved at a specific location in the juxtamembrane region 11 amino acids distal to the cell membrane, this cleavage has little sequence specificity. However, proline substitution at the P2' or P3' position or deletion of the epidermal growth factor (EGF) domain completely blocks the rapid phorbol ester-induced cleavage, but does not affect the slower basal proteolytic shedding. Insertion of the 15-residue membrane-proximal region (MPR) of L-selectin into the heterologous protein B7.2 results in a molecule that undergoes constitutive proteolytic turnover. In contrast, insertion of both the EGF domain and the MPR confers susceptibility to both slow constitutive shedding and the rapid proteolytic cleavage induced by phorbol 12-myristate 13-acetate. These results demonstrate that constitutive and induced L-selectin cleavage are separable processes and that the rapid phorbol ester-induced shedding requires the presence of the EGF domain, a sequence that is remote from the cleavage site.

T cells responding to antigen in vivo down-regulate L-selectin, the lymph node homing receptor, as they develop into activated effector cells. The concomitant up-regulation of the proinflammatory adhesion molecules LFA-1, CD44, and VLA-4 suggests that, after their release into the circulation, they traffic to sites of antigen deposition and inflammation. Previous evidence, however, has suggested a role for L-selectin in the recruitment of both neutrophils and lymphocytes into sites of inflammation, which would indicate that these L-selectin(-) effector cells could not be the precursors of inflammatory cells. We therefore directly tested whether L-selectin(-) T cells activated in vivo are capable of homing to model inflammatory sites. L-selectin(-) cells isolated from mice primed with alloantigen or with a contact sensitizer migrated to inflammation markedly better than L-selectin(+) cells from the same animals. Furthermore, the analogous population of CD44(hi)integrin(hi) cells from intravenously primed L-selectin knockout mice traffic efficiently to inflammatory sites and reject allogeneic skin grafts with normal kinetics. These data demonstrate that the previously described L-selectin(-) population of T cells that differentiate into effectors in spleen and lymph nodes subsequently traffic to inflammatory sites, due in part to their increased expression of other proinflammatory adhesion molecules.