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Evasion of innate immunity by parasitic protozoa.

Parasitic protozoa are a major cause of global infectious disease. These eukaryotic pathogens have evolved with the vertebrate immune system and typically produce long-lasting chronic infections. A critical step in their host interaction is the evasion of innate immune defenses. The ability to avoid attack by humoral effector mechanisms, such as complement lysis, is of particular importance to extracellular parasites, whereas intracellular protozoa must resist killing by lysosomal enzymes and toxic metabolites. They do so by remodeling the phagosomal compartments in which they reside and by interfering with signaling pathways that lead to cellular activation. In addition, there is growing evidence that protozoan pathogens modify the antigen-presenting and immunoregulatory functions of dendritic cells, a process that facilitates their evasion of both innate and adaptive immunity.

Animals↗

The adverse effect of pregnancy on macrophage activation.

Recently a remarkable decrease in the resistance to infection with Listeria monocytogenes and Toxoplasma gondii in pregnant mice was demonstrated. Since activated macrophages are important in the resistance against these organisms, studies were performed to determine whether pregnancy has an adverse effect on the activation of macrophages and effector functions of these macrophages. Peritoneal macrophages from normal pregnant and virgin mice or from pregnant and virgin mice previously injected with Corynebacterium parvum to activate peritoneal macrophages were challenged with EL-4 tumor cells to test for macrophage cytotoxicity or with T. gondii to test for the capacity to kill an intracellular pathogen. There was no consistent difference between the enhanced cytotoxic capacity of activated macrophages from pregnant and virgin mice. Macrophages from C. parvum-treated pregnant mice were deficient in their ability to kill or inhibit multiplication of T. gondii when compared to macrophages from C. parvum-treated control mice (P less than 0.001). Activated macrophages from pregnant mice were only able to mildly inhibit multiplication of T. gondii when compared to macrophages from control mice. Thus, there was a dissociation of effector function of activated macrophages from pregnant mice; they were defective in their ability to kill an intracellular pathogen but killed tumor target cells as well as did activated macrophages from virgin mice.

Animals↗

Construction and characterization of an effector strain of Streptococcus mutans for replacement therapy of dental caries.

An effector strain has been constructed for use in the replacement therapy of dental caries. Recombinant DNA methods were used to make the Streptococcus mutans supercolonizing strain, JH1140, lactate dehydrogenase deficient by deleting virtually all of the ldh open reading frame (ORF). To compensate for the resulting metabolic imbalance, a supplemental alcohol dehydrogenase activity was introduced by substituting the adhB ORF from Zymomonas mobilis in place of the deleted ldh ORF. The resulting clone, BCS3-L1, was found to produce no detectable lactic acid during growth on a variety of carbon sources, and it produced significantly less total acid due to its increased production of ethanol and acetoin. BCS3-L1 was significantly less cariogenic than JH1140 in both gnotobiotic- and conventional-rodent models. It colonized the teeth of conventional rats as well as JH1140 in both aggressive-displacement and preemptive-colonization models. No gross or microscopic abnormalities of major organs were associated with oral colonization of rats with BCS3-L1 for 6 months. Acid-producing revertants of BCS3-L1 were not observed in samples taken from infected animals (reversion frequency, <10(-3)) or by screening cultures grown in vitro, where no revertants were observed among 10(5) colonies examined on pH indicator medium. The reduced pathogenic potential of BCS3-L1, its strong colonization potential, and its genetic stability suggest that this strain is well suited to serve as an effector strain in the replacement therapy of dental caries in humans.

Animals↗

Neutrophils sustain pathogenic CD8+ T cell responses in the heart.

This study explores the influence of innate immunity on CD8(+) T-cell responses against heart tissue. Adoptive transfer of ovalbumin-specific CD8(+) effector T cells into CMy-mOva mice, which express ovalbumin in cardiac myocytes, results in a lethal acute myocarditis. The inflammatory infiltrate in the heart includes neutrophils as well as T cells. We used anti-Ly6G antibody to transiently deplete neutrophils at the time of onset of disease. By day 7 after receiving 5 x 10(5) CD8(+) effector T cells, 100% of control Ig-treated CMy-mOva mice had died, while 85% of anti-Ly6G-treated mice survived indefinitely. CD8(+) T-cell infiltration and tissue damage were present in both groups, but the disease was limited in the anti-Ly6G-treated mice, with a rapid disappearance of the adoptively transferred CD8(+) T cells within 11 days. Recovery occurred even though blood neutrophil counts began to rise 48 hours after the last anti-Ly6G treatment. Recovery was associated with a chronic CD4(+) cell infiltrate, and a rapid decline in expression of IFN-gamma and IP-10 mRNA in the myocardium. Neutrophil depletion did not effect survival of CMy-mOva mice that received 3 x 10(6) CD8(+) T cells. These data show that granulocytic inflammation sustains CD8(+) T-cell-mediated heart disease, which has important implications for the pathogenesis and treatment of acute myocarditis and allograft rejection.

Animals↗

A critical role for interleukin 18 in primary and memory effector responses to Listeria monocytogenes that extends beyond its effects on Interferon gamma production.

The stimulation of interferon (IFN)-gamma by interleukin (IL)-12 has been shown to provide protection from intracellular pathogens such as Listeria monocytogenes. Tumor necrosis factor (TNF) is also a major player in the resolution of Listeria infections and is suggested to have more global effects than can be explained by the induction of IFN-gamma alone. Since IL-18 synergizes with IL-12 to induce IFN-gamma production by natural killer and T helper (Th)1 cells, we determined its role in responses to Listeria. IL-18 appeared to be even more potent than either IL-12 or IFN-gamma for protection against this pathogen and IL-18 enhanced bacterial clearance in the complete absence of IFN-gamma. Indeed IL-18 was comparable to TNF in its ability to resolve the infection and showed a lowered protective capacity in the absence of TNF. Moreover, IL-18 induced macrophages to secrete both TNF and nitric oxide after a Listeria infection. IL-18 was also essential for optimal IFN-gamma production by antigen-specific T cells. Therefore, IL-18 operates via its effects on both the innate immune response, including macrophages, as well as on Th1 cells, to protect against Listeria.

Animals↗

A polytope DNA vaccine elicits multiple effector and memory CTL responses and protects against human papillomavirus 16 E7-expressing tumour.

Vaccine-induced CD8 T cells directed to tumour-specific antigens are recognised as important components of protective and therapeutic immunity against tumours. Where tumour antigens have pathogenic potential or where immunogenic epitopes are lost from tumours, development of subunit vaccines consisting of multiple individual epitopes is an attractive alternative to immunising with whole tumour antigen. In the present study we investigate the efficacy of two DNA-based multiepitope ('polytope') vaccines containing murine (H-2b) and human (HLA-A*0201)-restricted epitopes of the E7 oncoprotein of human papillomavirus type 16, in eliciting tumour-protective cytotoxic T-lymphocyte (CTL) responses. We show that the first of these polytopes elicited powerful effector CTL responses (measured by IFN-gamma ELISpot) and long-lived memory CTL responses (measured by functional CTL assay and tetramers) in immunised mice. The responses could be boosted by immunisation with a recombinant vaccinia virus expressing the polytope. Responses induced by immunisation with polytope DNA alone partially protected against infection with recombinant vaccinia virus expressing the polytope. Complete protection was afforded against challenge with an E7-expressing tumour, and reduced growth of nascent tumours was observed. A second polytope differing in the exact composition and order of CTL epitopes, and lacking an inserted endoplasmic reticulum targeting sequence and T-helper epitope, induced much poorer CTL responses and failed to protect against tumour challenge. These observations indicate the validity of a DNA polytope vaccine approach to human papillomavirus E7-associated carcinoma, and underscore the importance of design in polytope vaccine construction.

Amino Acid Sequence↗

Immunomodulation by a low-virulence, agerminative variant of Candida albicans. Further evidence for macrophage activation as one of the effector mechanisms of nonspecific anti-infectious protection.

Systemic infection of mice with a Candida albicans strain (PCA-2) incapable of yeast-mycelial conversion is known to activate host macrophages and confer protection against subsequent challenge with highly pathogenic cells of the same species or by other micro-organisms. In an attempt to define the relative contributions of different immune components to the protection mediated by PCA-2, we evaluated the effect of manipulations known to selectively deplete immune functions. By means of cytostatic drug or silica induced toxicity, it was possible to demonstrate that no crucial role in protection is played by cytotoxic T lymphocytes or B cells, nor by PCA-2 induced granulocytosis alone. The cells responsible for this effect were dacarbazine-resistant silica-sensitive macrophages whose activity in vivo paralleled the in vitro expression of splenic candidacidal activity. Macrophage activation by PCA-2 and increased anti-Candida resistance did not result from an immunological response mediated by T-dependent effectors, as these effects could be reproduced in athymic mice.

Adjuvants, Immunologic↗

Autophagy is a defense mechanism inhibiting BCG and Mycobacterium tuberculosis survival in infected macrophages.

Mycobacterium tuberculosis is an intracellular pathogen persisting within phagosomes through interference with phagolysosome biogenesis. Here we show that stimulation of autophagic pathways in macrophages causes mycobacterial phagosomes to mature into phagolysosomes. Physiological induction of autophagy or its pharmacological stimulation by rapamycin resulted in mycobacterial phagosome colocalization with the autophagy effector LC3, an elongation factor in autophagosome formation. Autophagy stimulation increased phagosomal colocalization with Beclin-1, a subunit of the phosphatidylinositol 3-kinase hVPS34, necessary for autophagy and a target for mycobacterial phagosome maturation arrest. Induction of autophagy suppressed intracellular survival of mycobacteria. IFN-gamma induced autophagy in macrophages, and so did transfection with LRG-47, an effector of IFN-gamma required for antimycobacterial action. These findings demonstrate that autophagic pathways can overcome the trafficking block imposed by M. tuberculosis. Autophagy, which is a hormonally, developmentally, and, as shown here, immunologically regulated process, represents an underappreciated innate defense mechanism for control of intracellular pathogens.

Animals↗

Immune function responds to selection for cuticular colour in Tenebrio molitor.

Cuticular colour in the mealworm beetle (Tenebrio molitor) is a quantitative trait, varying from tan to black. Population level variation in cuticular colour has been linked to pathogen resistance in this species and in several other insects: darker individuals are more resistant to pathogens. Given that cuticular colour has a heritable component, we have taken an experimental evolution approach: we selected 10 lines for black and 10 lines for tan adult cuticular phenotypes over at least six generations and measured the correlated responses to selection in a range of immune effector systems. Our results show that two immune parameters related to resistance (haemocyte density and pre-immune challenge activity of phenoloxidase (PO)) were significantly higher in selection lines of black beetles compared to tan lines. This may help to explain increased resistance to pathogens in darker individuals. Cuticular colour is dependent upon melanin production, which requires the enzyme PO that is present in its inactive form inside haemocytes. Thus, the observed correlated response to selection upon cuticular colour and immune variables probably results from these traits' shared dependence on melanin production.

Analysis of Variance↗

Toll-like receptor-4 signaling mediates pulmonary neutrophil sequestration in response to gram-positive bacterial enterotoxin.

BACKGROUND: Toll-like receptors (TLRs) serve as mediators of innate immune responses to pathogen-associated molecular patterns (PAMPs) which include lipopolysaccharide (LPS) and staphylococcal enterotoxin B (SEB). TLR-4 is thought to act as the primary effector of LPS recognition and TLR-2 is thought to mediate responses to Gram-positive bacterial proteins. Chemokines such as macrophage inflammatory protein (MIP-2) are peptides that are responsible for lung neutrophil (PMN) sequestration following an infectious or inflammatory insult. Given the Gram-positive origin of SEB, we hypothesized that mice with altered TLR-4 signaling would exhibit no difference in lung PMN sequestration following SEB when compared to wild-type mice. METHODS: Wild-type and TLR-4 mutant mice were administered intratracheal saline, LPS (Escherichia coli 0.1 mg/kg), or SEB (1 mg/kg). After 24 h, lung PMN accumulation was determined by myeloperoxidase (MPO) assay and bronchoalveolar lavage fluid cell count (BALfcc). Total lung and BALf MIP-2 was measured by enzyme-linked immunosorbent assay. RESULTS: There was an increase in lung PMN accumulation (by both MPO and BALfcc) and MIP-2 following LPS and SEB in wild-type mice compared to saline-treated controls. In contrast, TLR-4 mice failed to exhibit an increase in lung MIP-2 or PMN accumulation following either LPS or SEB compared to wild-type mice. CONCLUSIONS: TLR-4 mutant mice are unresponsive to intratracheal LPS. SEB elicited an increase in lung MIP-2 and PMN accumulation in wild-type mice. However, TLR-4 mutant mice were protected from this process. This suggests that TLR-4 signaling may mediate the responses to other PAMPs in addition to LPS.

Animals↗

Natural killer cells as an initial defense against pathogens.

Natural killer (NK) cells serve as a crucial first line of defense against tumors and a diverse range of pathogens. Recognition of infection by NK cells is accomplished by the activation of receptors on the NK cell surface, which initiate NK cell effector functions. Many of the receptors and ligands involved in NK cell antimicrobial activity have been identified, and we are beginning to appreciate how they function during infection. In addition, NK cells are activated by cytokines (e.g. interleukin 12 and type I interferons), which are products of activated macrophages and dendritic cells. In response to these activating stimuli, NK cells secrete cytokines and chemokines and lyse target cells. Recent studies have focused on the mechanisms by which NK cells recognize and respond to viruses, parasites and bacteria, and on the unique role of NK cells in innate immunity to infection.

Animals↗

Experimental evolution reveals genetic routes for adaptive loss of the antibacterial type VI secretion system.

The type VI secretion system (T6SS) is a contractile nanomachine used by Gram-negative bacteria to deliver effector proteins into target cells, contributing to both interbacterial competition and pathogenesis. Although T6SS gene clusters are present in recently isolated commensal and pathogenic Escherichia coli strains, they are absent from classical laboratory strains that have been propagated for decades in pure cultures, suggesting that T6SS can be lost in the absence of competition. Here, we combined experimental evolution with whole-genome sequencing to track the fate of the enteroaggregative Escherichia coli (EAEC) Sci1 T6SS during competition with either T6SS-susceptible or T6SS-immune bacteria. After &#x223c;640 generations, T6SS activity was largely maintained during competition with T6SS-susceptible bacteria, whereas &#x223c;90% of clones evolved with T6SS-immune bacteria lost or attenuated T6SS activity through diverse mutations within the sci1 promoter, essential T6SS structural genes, or the rfaH transcriptional antiterminator. We identified two RfaH-binding ops elements within the sci1 cluster, revealing antitermination as a regulatory element of EAEC T6SS transcription, which is conserved among Enterobacteriaceae. Our findings highlight how experimental evolution can reveal the selective forces shaping T6SS maintenance and identify new regulatory components controlling its activity.

Journal Article↗

Cysteine proteases in phytopathogenic bacteria: identification of plant targets and activation of innate immunity.

Phytopathogenic bacteria use the type-III secretion system (TTSS) to inject effector proteins into plant cells, presumably to colonize their hosts. The function of these proteins inside plant cells has remained a mystery for years. The recent discovery that the effectors XopD, AvrXv4, AvrPphB, and AvrRpt2 have cysteine protease functions reveals that the proteolysis of host substrates is an important strategy employed by pathogens to alter plant physiology. Moreover, the characterization of these proteases and their targets provides new insight to mechanisms of bacterial virulence and the activation of plant immunity.

Bacteria↗

Phenotype and function of intestinal dendritic cells.

It is now appreciated that dendritic cells (DCs) play a primary role in oral tolerance and defense against mucosal pathogens. Specific DC subpopulations are localized to discrete regions within primary inductive tissues, like the Peyer's patch and mesenteric lymph node, and effector sites, like the lamina propria, and may have unique roles in driving regulatory, effector and memory T cell responses. Certain DC subpopulations may also help maintain T cell responses at sites of abnormal intestinal inflammation. While early in our understanding, knowledge about the involvement of DC subpopulations in the regulation of mucosal immunity may well provide a basis for the development of novel vaccines and therapeutics.

Animals↗

RAC/ROP GTPases: 'hubs' for signal integration and diversification in plants.

RAC/ROP GTPases are a family of plant-specific signaling molecules solely representing the Ras and Rho family of Ras-related G proteins in plants. RAC/ROPs potentially interact with cell surface-associated signal perception apparatus for a broad range of extracellular stimuli, including hormones, pathogen elicitors and abiotic stress, and mediate diverse cellular pathways in response to these signals. They are also known to interact with multiple effectors, affecting cellular and biochemical systems that regulate actin dynamics, reactive oxygen species production, proteolysis, and gene expression. RAC/ROPs are, thus, ideally suited as integrators for multiple signals and as coordinators of diverse cellular pathways to control growth, differentiation, development and defense responses. Recent findings that suggest how RAC/ROP signaling activity is regulated and how functional specificity can be achieved are discussed here.

Arabidopsis Proteins↗

Initiation of RPS2-specified disease resistance in Arabidopsis is coupled to the AvrRpt2-directed elimination of RIN4.

Plants have evolved a sophisticated innate immune system to recognize invading pathogens and to induce a set of host defense mechanisms resulting in disease resistance. Pathogen recognition is often mediated by plant disease resistance (R) proteins that respond specifically to one or a few pathogen-derived molecules. This specificity has led to suggestions of a receptor-ligand mode of R protein function. Delivery of the bacterial effector protein AvrRpt2 by Pseudomonas syringae specifically induces disease resistance in Arabidopsis plants expressing the RPS2 R protein. We demonstrate that RPS2 physically interacts with Arabidopsis RIN4 and that AvrRpt2 causes the elimination of RIN4 during activation of the RPS2 pathway. AvrRpt2-mediated RIN4 elimination also occurs in the rps2, ndr1, and Atrar1 mutant backgrounds, demonstrating that this activity can be achieved independent of an RPS2-mediated signaling pathway. Therefore, we suggest that RPS2 initiates signaling based upon perception of RIN4 disappearance rather than direct recognition of AvrRpt2.

Alleles↗

Early nutrition and the development of immune function in the neonate.

The present review will concentrate on the development of the gut-associated lymphoid tissue and the role of early nutrition in promoting immune function. The intestine is the largest immune organ in the body, and as such is the location for the majority of lymphocytes and other immune effector cells. The intestine is exposed to vast quantities of dietary and microbial antigens, and is the most common portal of entry for pathogens, some of which are potentially lethal. The development of normal immune function of the intestine is therefore vital for survival, and is dependent on appropriate antigen exposure and processing, and also an intact intestinal barrier. In early life innate mechanisms of defence are probably more important than active or adaptive mechanisms in responding to an infectious challenge, since the healthy neonate is immunologically naïve (has not seen antigen) and has not acquired immunological memory. During this period maternal colostrum and milk can significantly augment resistance to enteric infections. The mechanisms of enhancing disease resistance are thought to be passive, involving a direct supply of anti-microbial factors, and active, by promoting the development of specific immune function. A tolerance response to dietary and non-invasive antigens is generally induced in the gut. However, it must also be able to mount an adequate immune response to ensure clearance of foreign antigens. It is now recognized that regulation of tolerance and active immune responses is critical to health, and failure to regulate these responses can lead to recurrent infections, inflammatory diseases and allergies. The education of the immune system in early life is thought to be critical in minimizing the occurrence of these immune-based disorders. During this phase of development maternal milk provides signals to the immune system that generate appropriate response and memory. One factor that has been proposed to contribute to the increase in the incidence of immune-based disorders, e.g. atopic diseases in Western countries, is thought to be the increased prevalence of formula-feeding.

Animals↗

Protection against infections by oral lactoferrin: evaluation in animal models.

It has been reported previously that oral administration of lactoferrin (LF) provides some host-protective effects against infections, cancers, and inflammations. In this review, we focus on the effect of oral LF on various infectious diseases and discuss the mechanism as elucidated in animal models. In the case of infections occurring at sites other than the digestive canal, it is unclear whether oral LF is absorbed from the intestine and exerts its protective effect at the site of infection. In preterm human infants, neonatal pigs, and rats with colitis, it was reported that LF is detectable in various body fluids after oral administration. We could not detect the transport of oral bovine LF into the blood of adult rats without gastrointestinal illness using several techniques, suggesting that there is an extremely low level of transport of LF, if any. Orally administered LF may act at the oro-gastro-intestinal mucosa and aid the defense system against infections through a network of mucosal immunity and systemic immunity. Indeed, it is reported that oral LF increases the number of cells in the leukocyte subset and cytokine (IFN-gamma and IL-18) production in the intestinal mucosa of mice. Regarding systemic immunity, we have observed an increase of leukocyte number, cytokine (IFN-gamma, TNF-alpha, IL-12, and IL-18) production, and effector activity of macrophages in response to LF administration in several animal models. These enhanced immune responses may contribute to eradication of the pathogen, resolution of the symptoms, and maintenance of the homeostasis during infectious diseases.

Administration, Oral↗