Neonatal Host Defense against Staphylococcal Infections

Preterm infants are especially susceptible to late-onset sepsis that is often due to Gram-positive bacterial infections resulting in substantial morbidity and mortality. Herein, we will describe neonatal innate immunity to Staphylococcus spp. comparing differences between preterm and full-term newborns with adults. Newborn innate immunity is distinct demonstrating diminished skin integrity, impaired Th1-polarizing responses, low complement levels, and diminished expression of plasma antimicrobial proteins and peptides, especially in preterm newborns. Characterization of distinct aspects of the neonatal immune response is defining novel approaches to enhance host defense to prevent and/or treat staphylococcal infection in this vulnerable population

A Neonatal Model of Intravenous Staphylococcus epidermidis Infection in Mice <24 h Old Enables Characterization of Early Innate Immune Responses

Staphylococcus epidermidis (SE) causes late onset sepsis and significant morbidity in catheterized preterm newborns. Animal models of SE infection are useful in characterizing disease mechanisms and are an important approach to developing improved diagnostics and therapeutics. Current murine models of neonatal bacterial infection employ intraperitoneal or subcutaneous routes at several days of age, and may, therefore, not accurately reflect distinct features of innate immune responses to bacteremia. In this study we developed, validated, and characterized a murine model of intravenous (IV) infection in neonatal mice <24 hours (h) old to describe the early innate immune response to SE. C57BL/6 mice <24 h old were injected IV with 106, 107, 108 colony-forming units (CFU) of SE 1457, a clinical isolate from a central catheter infection. A prospective injection scoring system was developed and validated, with only high quality injections analyzed. Newborn mice were euthanized between 2 and 48 h post-injection and spleen, liver, and blood collected to assess bacterial viability, gene expression, and cytokine production. High quality IV injections demonstrated inoculum-dependent infection of spleen, liver and blood. Within 2 h of injection, SE induced selective transcription of TLR2 and MyD88 in the liver, and increased systemic production of plasma IL-6 and TNF-α. Despite clearance of bacteremia and solid organ infection within 48 h, inoculum-dependent impairment in weight gain was noted. We conclude that a model of IV SE infection in neonatal mice <24 h old is feasible, demonstrating inoculum-dependent infection of solid organs and a pattern of bacteremia, rapid and selective innate immune activation, and impairment of weight gain typical of infected human neonates. This novel model can now be used to characterize immune ontogeny, evaluate infection biomarkers, and assess preventative and therapeutic modalities

TLR2 Mediates Recognition of Live Staphylococcus epidermidis and Clearance of Bacteremia

Background: Staphylococcus epidermidis (SE) is a nosocomial pathogen that causes catheter-associated bacteremia in the immunocompromised, including those at the extremes of age, motivating study of host clearance mechanisms. SE-derived soluble components engage TLR2; but additional signaling pathways have also been implicated, and TLR2 can play complex, at times detrimental, roles in host defense against other Staphylococcal spp. The role of TLR2 in responses of primary blood leukocytes to live SE and in clearance of SE bacteremia, the most common clinical manifestation of SE infection, is unknown. Methodology/Principal Findings: We studied TLR2-mediated recognition of live clinical SE strain 1457 employing TLR2- transfected cells, neutralizing anti-TLR antibodies and TLR2-deficient mice. TLR2 mediated SE-induced cytokine production in human embryonic kidney cells, human whole blood and murine primary macrophages, in part via recognition of a soluble TLR2 agonist. After i.v. challenge with SE, early (1 h) cytokine/chemokine production and subsequent clearance of bacteremia (24-48 h) were markedly impaired in TLR2-deficient mice. Conclusions/Significance: TLR2 mediates recognition of live SE and clearance of SE bacteremia in vivo

Investigating Potential Applications of the Fish Anti-Microbial Peptide Pleurocidin: A Systematic Review

The anti-microbial peptide (AMP) pleurocidin is found in winter flounder (Pseudopleuronectes americanus), an Atlantic flounder species. There is promising evidence for clinical, aquaculture, and veterinary applications of pleurocidin. This review provides an overview of the current literature available on pleurocidin to guide future research directions. By fully elucidating pleurocidin’s mechanism of action and developing novel treatments against pathogenic microbes, populations of flatfish and humans can be protected. This review consulted publications from PubMed and Environment Complete with search terms such as “pleurocidin”, “winter flounder”, and “antimicrobial”. The fish immune system includes AMPs as a component of the innate immune system. Pleurocidin, one of these AMPs, has been found to be effective against various Gram-positive and Gram-negative bacteria. More investigations are required to determine pleurocidin’s suitability as a treatment against antibiotic-resistant pathogens. There is promising evidence for pleurocidin as a novel anti-cancer therapy. The peptide has been found to display potent anti-cancer effects against human cancer cells. Research efforts focused on pleurocidin may result in novel treatment strategies against antibiotic-resistant bacteria and cancer. More research is required to determine if the peptide is a suitable candidate to be developed into a novel anti-microbial treatment. Some of the microbes susceptible to the peptide are also pathogens of fish, suggesting its suitability as a therapeutic treatment for fish species

Review: Examining the Natural Role of Amphibian Antimicrobial Peptide Magainin

Host defense peptides (HDPs) are a group of antimicrobial peptides (AMPs) that are crucial components of the innate immune system of many different organisms. These small peptides actively kill microbes and prevent infection. Despite the presence of AMPs in the amphibian immune system, populations of these organisms are in decline globally. Magainin is an AMP derived from the African clawed frog (Xenopus laevis) and has displayed potent antimicrobial effects against a wide variety of microbes. Included in this group of microbes are known pathogens of the African clawed frog and other amphibian species. Arguably, the most deleterious amphibious pathogen is Batrachochytrium dendrobatidis, a chytrid fungus. Investigating the mechanism of action of magainin can help understand how to effectively fight off infection. By understanding amphibian AMPs&rsquo; role in the frog, a potential conservation strategy can be developed for other species of amphibians that are susceptible to infections, such as the North American green frog (Rana clamitans). Considering that population declines of these organisms are occurring globally, this effort is crucial to protect not only these organisms but the ecosystems they inhabit as well

Neonatal Host Defense against Staphylococcal Infections

Preterm infants are especially susceptible to late-onset sepsis that is often due to Gram-positive bacterial infections resulting in substantial morbidity and mortality. Herein, we will describe neonatal innate immunity to Staphylococcus spp. comparing differences between preterm and full-term newborns with adults. Newborn innate immunity is distinct demonstrating diminished skin integrity, impaired Th1-polarizing responses, low complement levels, and diminished expression of plasma antimicrobial proteins and peptides, especially in preterm newborns. Characterization of distinct aspects of the neonatal immune response is defining novel approaches to enhance host defense to prevent and/or treat staphylococcal infection in this vulnerable population

4T1 Murine Mammary Carcinoma Cells Enhance Macrophage-Mediated Innate Inflammatory Responses.

Tumor progression and the immune response are intricately linked. While it is known that cancers alter macrophage inflammatory responses to promote tumor progression, little is known regarding how cancers affect macrophage-dependent innate host defense. In this study, murine bone-marrow-derived macrophages (BMDM) were exposed to murine carcinoma-conditioned media prior to assessment of the macrophage inflammatory response. BMDMs exposed to 4T1 mammary carcinoma-conditioned medium demonstrated enhanced production of pro-inflammatory cytokines tumor necrosis factor α, interleukin-6, and CCL2 in response to lipopolysaccharide (LPS) while production of interleukin-10 remained unchanged. The increased LPS-induced production of pro-inflammatory cytokines was transient and correlated with enhanced cytokine production in response to other Toll-like receptor agonists, including peptidoglycan and flagellin. In addition, 4T1-conditioned BMDMs exhibited strengthened LPS-induced nitric oxide production and enhanced phagocytosis of Escherichia coli. 4T1-mediated augmentation of macrophage responses to LPS was partially dependent on the NFκB pathway, macrophage-colony stimulating factor, and actin polymerization, as well as the presence of 4T1-secreted extracellular vesicles. Furthermore, peritoneal macrophages obtained from 4T1 tumor-bearing mice displayed enhanced pro-inflammatory cytokine production in response to LPS. These results suggest that uptake of 4T1-secreted factors and actin-mediated ingestion of 4T1-secreted exosomes by macrophages cause a transient enhancement of innate inflammatory responses. Mammary carcinoma-mediated regulation of innate immunity may have significant implications for our understanding of host defense and cancer progression

4T1-conditioned macrophages exhibit increased phagocytosis of <i>E</i>. <i>coli</i> bioparticles.

<p>BMDMs were treated for 24 h with 4T1- or HC11-conditioned medium (CM). BMDMs were then washed with PBS and incubated for 2 h with <i>E</i>. <i>coli</i> bioparticles, which fluoresce only under acidic conditions present in endosomes. <i>E</i>. <i>coli</i> phagocytosis was determined by measuring BMDM fluorescence by flow cytometry. As a negative control, BMDMs were pre-treated for 1 h with actin polymerization inhibitor cytochalasin D (10 μM) prior to the addition of <i>E</i>. <i>coli</i> bioparticles. Phagocytic activity was normalized to the uptake of <i>E</i>. <i>coli</i> bioparticles by DMDMs treated with DMEM alone. Results are shown as the mean (± SEM) of at least 4 independent experiments. Statistical analyses were done by ANOVA, followed by Tukey’s multiple comparison tests; * p < 0.05.</p

Peritoneal macrophages from 4T1 tumor-bearing mice demonstrate enhanced sensitivity to LPS.

<p>Peritoneal macrophages from control (n = 4) or 4T1-bearing mice (n = 6) were stimulated with 100 ng/ml LPS for 4 h. Culture supernatants were collected and assessed for the levels of cytokines by ELISA. Statistical analyses were done by ANOVA, followed by Tukey’s multiple comparison tests; * p < 0.05.</p