A scabies mite serpin interferes with complement-mediated neutrophil functions and promotes staphylococcal growth.

PubMed

Swe, Pearl M; Fischer, Katja

2014-06-01

Scabies is a contagious skin disease caused by the parasitic mite Sarcoptes scabiei. The disease is highly prevalent worldwide and known to predispose to secondary bacterial infections, in particular by Streptococcus pyogenes and Staphylococcus aureus. Reports of scabies patients co-infected with methicillin resistant S. aureus (MRSA) pose a major concern for serious down-stream complications. We previously reported that a range of complement inhibitors secreted by the mites promoted the growth of S. pyogenes. Here, we show that a recently characterized mite serine protease inhibitor (SMSB4) inhibits the complement-mediated blood killing of S. aureus. Blood killing of S. aureus was measured in whole blood bactericidal assays, counting viable bacteria recovered after treatment in fresh blood containing active complement and phagocytes, treated with recombinant SMSB4. SMSB4 inhibited the blood killing of various strains of S. aureus including methicillin-resistant and methicillin-sensitive isolates. Staphylococcal growth was promoted in a dose-dependent manner. We investigated the effect of SMSB4 on the complement-mediated neutrophil functions, namely phagocytosis, opsonization and anaphylatoxin release, by flow cytometry and in enzyme linked immuno sorbent assays (ELISA). SMSB4 reduced phagocytosis of S. aureus by neutrophils. It inhibited the deposition of C3b, C4b and properdin on the bacteria surface, but did not affect the depositions of C1q and MBL. SMSB4 also inhibited C5 cleavage as indicated by a reduced C5b-9 deposition. We postulate that SMSB4 interferes with the activation of all three complement pathways by reducing the amount of C3 convertase formed. We conclude that SMSB4 interferes with the complement-dependent killing function of neutrophils, thereby reducing opsonization, phagocytosis and further recruitment of neutrophils to the site of infection. As a consequence secreted scabies mites complement inhibitors, such as SMSB4, provide favorable conditions for the onset of S. aureus co-infection in the scabies-infected microenvironment by suppressing the immediate host immune response.

Functional Activity of Antibodies Directed towards Flagellin Proteins of Non-Typhoidal Salmonella

PubMed Central

Boyd, Mary A.; Wang, Jin Y.; Tulapurkar, Mohan E.; Pasetti, Marcela F.; Levine, Myron M.; Simon, Raphael

2016-01-01

Non-typhoidal Salmonella (NTS) serovars Typhimurium and Enteritidis are major causes of invasive bacterial infections in children under 5 years old in sub-Saharan Africa, with case fatality rates of ~20%. There are no licensed NTS vaccines for humans. Vaccines that induce antibodies against a Salmonella Typhi surface antigen, Vi polysaccharide, significantly protect humans against typhoid fever, establishing that immune responses to Salmonella surface antigens can be protective. Flagella proteins, abundant surface antigens in Salmonella serovars that cause human disease, are also powerful immunogens, but the functional capacity of elicited anti-flagellar antibodies and their role in facilitating bacterial clearance has been unclear. We examined the ability of anti-flagellar antibodies to mediate microbial killing by immune system components in-vitro and assessed their role in protecting mice against invasive Salmonella infection. Polyclonal (hyperimmune sera) and monoclonal antibodies raised against phase 1 flagellin proteins of S. Enteritidis and S. Typhimurium facilitated bacterial uptake and killing of the homologous serovar pathogen by phagocytes. Polyclonal anti-flagellar antibodies accompanied by complement also achieved direct bacterial killing. Serum bactericidal activity was restricted to Salmonella serovars expressing the same flagellin used as immunogen. Notably, individual anti-flagellin monoclonal antibodies with complement were not bactericidal, but this biological activity was restored when different monoclonal anti-flagellin antibodies were combined. Passive transfer immunization with a monoclonal IgG antibody specific for phase 1 flagellin from S. Typhimurium protected mice against lethal challenge with a representative African invasive S. Typhimurium strain. These findings have relevance for the use of flagellin proteins in NTS vaccines, and confirm the role of anti-flagellin antibodies as mediators of protective immunity. PMID:26998925

Immune functions of the garment workers.

PubMed

Sultana, R; Ferdous, K J; Hossain, M; Zahid, M S H; Islam, L N

2012-10-01

Occupational exposure to cotton dust, fibers, metal fumes and different chemicals used in the aparrel manufacturing industries cause a wide range of physical and psychological health problems in the garment workers that may also affect their immune function. To assess the immune system function in garment workers. A total of 45 workers of a garment factory, and 41 control subjects, not exposed to the garment working environment were enrolled in this study. In the study subjects, the complement system function was assessed as bactericidal activity on Escherichia coli DH5α cells using the standard plate count method. Serum complement components C3 and C4 were measured by immunoprecipitation, and IgG was measured by immunonephelometry. The bactericidal activity of serum complement in the garment workers (range: 93.5%-99.9%) was significantly (p<0.01) lower than that in the controls (range: 98.6%-100%). The heat-inactivated serum of the workers showed a significantly enhanced bactericidal activity. In the garment workers, the mean levels of complement C3, and C4 were 1.75 and 0.26 g/L, respectively that were close to those of the controls. The mean IgG level in the garment workers was 13.5 g/L that was significantly (p<0.001) higher than that in the controls. Working in a garment factory may affect the immune system.

Characterization of diverse subvariants of the meningococcal factor H (fH) binding protein for their ability to bind fH, to mediate serum resistance, and to induce bactericidal antibodies.

PubMed

Seib, Kate L; Brunelli, Brunella; Brogioni, Barbara; Palumbo, Emmanuelle; Bambini, Stefania; Muzzi, Alessandro; DiMarcello, Federica; Marchi, Sara; van der Ende, Arie; Aricó, Beatrice; Savino, Silvana; Scarselli, Maria; Comanducci, Maurizio; Rappuoli, Rino; Giuliani, Marzia M; Pizza, Mariagrazia

2011-02-01

Neisseria meningitidis is a commensal of the human nasopharynx but is also a major cause of septicemia and meningitis. The meningococcal factor H binding protein (fHbp) binds human factor H (fH), enabling downregulation of complement activation on the bacterial surface. fHbp is a component of two serogroup B meningococcal vaccines currently in clinical development. Here we characterize 12 fHbp subvariants for their level of surface exposure and ability to bind fH, to mediate serum resistance, and to induce bactericidal antibodies. Flow cytometry and Western analysis revealed that all strains examined expressed fHbp on their surface to different extents and bound fH in an fHbp-dependent manner. However, differences in fH binding did not always correlate with the level of fHbp expression, indicating that this is not the only factor affecting the amount of fH bound. To overcome the issue of strain variability in fHbp expression, the MC58ΔfHbp strain was genetically engineered to express different subvariants from a constitutive heterologous promoter. These recombinant strains were characterized for fH binding, and the data confirmed that each subvariant binds different levels of fH. Surface plasmon resonance revealed differences in the stability of the fHbp-fH complexes that ranged over 2 orders of magnitude, indicating that differences in residues between and within variant groups can influence fH binding. Interestingly, the level of survival in human sera of recombinant MC58 strains expressing diverse subvariants did not correlate with the level of fH binding, suggesting that the interaction of fHbp with fH is not the only function of fHbp that influences serum resistance. Furthermore, cross-reactive bactericidal activity was seen within each variant group, although the degree of activity varied, suggesting that amino acid differences within each variant group influence the bactericidal antibody response.

Detection and characterisation of Complement protein activity in bovine milk by bactericidal sequestration assay.

PubMed

Maye, Susan; Stanton, Catherine; Fitzgerald, Gerald F; Kelly, Philip M

2015-08-01

While the Complement protein system in human milk is well characterised, there is little information on its presence and activity in bovine milk. Complement forms part of the innate immune system, hence the importance of its contribution during milk ingestion to the overall defences of the neonate. A bactericidal sequestration assay, featuring a Complement sensitive strain, Escherichia coli 0111, originally used to characterise Complement activity in human milk was successfully applied to freshly drawn bovine milk samples, thus, providing an opportunity to compare Complement activities in both human and bovine milks. Although not identical in response, the levels of Complement activity in bovine milk were found to be closely comparable with that of human milk. Differential counts of Esch. coli 0111 after 2 h incubation were 6.20 and 6.06 log CFU/ml, for raw bovine and human milks, respectively - the lower value representing a stronger Complement response. Exposing bovine milk to a range of thermal treatments e.g. 42, 45, 65, 72, 85 or 95 °C for 10 min, progressively inhibited Complement activity by increasing temperature, thus confirming the heat labile nature of this immune protein system. Low level Complement activity was found, however, in 65 and 72 °C heat treated samples and in retailed pasteurised milk which highlights the outer limit to which high temperature, short time (HTST) industrial thermal processes should be applied if retention of activity is a priority. Concentration of Complement in the fat phase was evident following cream separation, and this was also reflected in the further loss of activity recorded in low fat variants of retailed pasteurised milk. Laboratory-based churning of the cream during simulated buttermaking generated an aqueous (buttermilk) phase with higher levels of Complement activity than the fat phase, thus pointing to a likely association with the milk fat globule membrane (MFGM) layer.

Inhibition of C5a-induced inflammation with preserved C5b-9-mediated bactericidal activity in a human whole blood model of meningococcal sepsis.

PubMed

Sprong, Tom; Brandtzaeg, Petter; Fung, Michael; Pharo, Anne M; Høiby, E Arne; Michaelsen, Terje E; Aase, Audun; van der Meer, Jos W M; van Deuren, Marcel; Mollnes, Tom E

2003-11-15

The complement system plays an important role in the initial defense against Neisseria meningitidis. In contrast, uncontrolled activation in meningococcal sepsis contributes to the development of tissue damage and shock. In a novel human whole blood model of meningococcal sepsis, we studied the effect of complement inhibition on inflammation and bacterial killing. Monoclonal antibodies (mAbs) blocking lectin and alternative pathways inhibited complement activation by N meningitidis and oxidative burst induced in granulocytes and monocytes. Oxidative burst was critically dependent on CD11b/CD18 (CR3) expression but not on Fc gamma-receptors. Specific inhibition of C5a using mAb 137-26 binding the C5a moiety of C5 before cleavage prohibited CR3 up-regulation, phagocytosis, and oxidative burst but had no effect on C5b-9 (TCC) formation, lysis, and bacterial killing. An mAb-blocking cleavage of C5, preventing C5a and TCC formation, showed the same effect on CR3, phagocytosis, and oxidative burst as the anti-C5a mAb but additionally inhibited TCC formation, lysis, and bacterial killing, consistent with a C5b-9-dependent killing mechanism. In conclusion, the anti-C5a mAb 137-26 inhibits the potentially harmful effects of N meningitidis-induced C5a formation while preserving complement-mediated bacterial killing. We suggest that this may be an attractive approach for the treatment of meningococcal sepsis.

Burn wound sepsis may be promoted by a failure of local antibacterial host defenses.

PubMed Central

Deitch, E A; Bridges, R M; Dobke, M; McDonald, J C

1987-01-01

Little attention has been focused on the local burn wound environment, even though burn wound sepsis is a common cause of death in the burn victim. To characterize the effect of the local burn wound environment on neutrophil function and metabolism, the opsonic activity of blister fluid specimens against Pseudomonas aeruginosa was measured as was the effect of blister fluid on control neutrophil oxygen consumption using preopsonized zymosan and f-met-leu-phe (FMLP) as stimuli. Blister fluid did not support the killing of P. aeruginosa by normal neutrophils as well as normal serum. Additionally, blister fluid inhibited zymosan-stimulated, but not FMLP-stimulated, neutrophil oxygen consumption. The inhibitory effect of blister fluid on zymosan-stimulated oxygen consumption correlated with the extent of complement activation, measured as C3d or C3AI (p less than 0.01). That blister fluid did not inhibit the FMLP-mediated respiratory burst supports the concept that the blister fluid inhibitory effect on the zymosan-mediated respiratory burst was mediated through the complement receptor. These findings that blister fluid can affect the bactericidal and metabolic activity of normal neutrophils support the concept that cellular function can be altered by the microenvironment in which the cells are bathed. This potential impairment of host defenses within the burn wound could predispose the burn victim to burn wound sepsis. PMID:3115207

DrsG from Streptococcus dysgalactiae subsp. equisimilis Inhibits the Antimicrobial Peptide LL-37

PubMed Central

Smyth, Danielle; Cameron, Ainslie; Davies, Mark R.; McNeilly, Celia; Hafner, Louise; Sriprakash, Kadaba S.

2014-01-01

SIC and DRS are related proteins present in only 4 of the >200 Streptococcus pyogenes emm types. These proteins inhibit complement-mediated lysis and/or the activity of certain antimicrobial peptides (AMPs). A gene encoding a homologue of these proteins, herein called DrsG, has been identified in the related bacterium Streptococcus dysgalactiae subsp. equisimilis. Here we show that geographically dispersed isolates representing 14 of 50 emm types examined possess variants of drsG. However, not all isolates within the drsG-positive emm types possess the gene. Sequence comparisons also revealed a high degree of conservation in different S. dysgalactiae subsp. equisimilis emm types. To examine the biological activity of DrsG, recombinant versions of two major DrsG variants, DrsGS and DrsGL, were expressed and purified. Western blot analysis using antisera raised to these proteins demonstrated both variants to be expressed and secreted into culture supernatants. Unlike SIC, but similar to DRS, DrsG does not inhibit complement-mediated lysis. However, like both SIC and DRS, DrsG is a ligand of the cathelicidin LL-37 and is inhibitory to its bactericidal activity in in vitro assays. Conservation of prolines in the C-terminal region also suggests that these residues are important in the biology of this family of proteins. This is the first report demonstrating the activity of an AMP-inhibitory protein in S. dysgalactiae subsp. equisimilis and suggests that inhibition of AMP activity is the primary function of this family of proteins. The acquisition of the complement-inhibitory activity of SIC may reflect its continuing evolution. PMID:24664506

Inhibition of intracellular bacterial replication in fibroblasts is dependent on the perforin-like protein (Perforin-2) encoded by macrophage expressed gene 1

PubMed Central

McCormack, Ryan; de Armas, Lesley R.; Shiratsuchi, Motoaki; Ramos, Jay; Podack, Eckhard R.

2013-01-01

Fibroblasts are known to eliminate intracellular bacteria, but the lethal hit of the bactericidal mechanism has not been defined. We show that primary embryonic and established fibroblasts can be induced by interferons or by intracellular bacterial infection to express a perforin-like mRNA previously described as macrophage expressed gene 1 (mpeg1). The presence and level of the perforin-like mRNA correlate with the ability of primary mouse embryonic fibroblasts (MEF) to eliminate intracellular bacteria. In addition, siRNA knock-down of the perforin-like molecule abolishes bactericidal activity and allows intracellular bacterial replication. Complementation of MEF in which the endogenous perforin-like molecule has been knocked down with an RFP-tagged version restores bactericidal activity. The perforin-like molecule has broad bactericidal specificity for pathogenic and non-pathogenic bacteria including Gram positive, Gram negative and acid fast bacteria. The perforin-like molecule renders previously lysozyme-resistant bacteria sensitive to lysis by lysozyme suggesting physical damage of the outer cell wall by the perforin-like protein. MEFs damage cell walls of intracellular bacteria by insertion, polymerization and pore-formation of the perforin-like protein, analogous to pore-formers of complement and Perforin-1 of cytolytic lymphocytes. We propose the name Perforin-2. PMID:23257510

Immune modulation following immunization with polyvalent vaccines in dogs.

PubMed

Strasser, Alois; May, Bettina; Teltscher, Andrea; Wistrela, Eva; Niedermüller, Hans

2003-08-15

A decline in T-cell-mediated immunity and transient state of immunosuppression after immunization has been reported in dogs. Nevertheless, dogs are still routinely vaccinated with polyvalent live vaccines and severe disease does not generally occur. In order to investigate these effects on the canine immune system and to elucidate possible mechanisms we determined the following immune parameters in the blood of 33 clinically sound German shepherd dogs before and after standard vaccination with a polyvalent vaccine against distemper, parvovirus, viral hepatitis, leptospirosis, kennel cough and rabies: white and differential blood cell count, the serum concentrations and/or activities of IL-1, IL-2, IFN-gamma, TNF-alpha, neopterin and IgG, natural killer (NK) cell activity, bactericidal activity and complement hemolytic activity, lymphocyte proliferation test (LPT) and nitroblue tetrazolium test (NBT). Our major findings were that significant postvaccinal decreases in T-cell mitogenic response to PHA and in neutrophil function and neopterin serum concentration were accompanied by simultaneous increase in plasma IgG and hemolytic complement activity. This suggests a transient shift in the balance between cell-mediated and humoral (T(H)1/T(H)2) immunity rather than immunosuppression. These results do not imply that dogs should not receive live vaccines, as the response to vaccines just seems to create a state of altered homeostasis when immunization elicits protection by humoral and cell-mediated immunity. However, these recognized compromises of immune function should be considered and vaccines still be applied only in healthy animals and strictly according to the rules and regulations given by the manufacturer.

Re-examination of the Neisser—Wechsberg (antibody prozone) phenomenon

PubMed Central

Muschel, L. H.; Gustafson, Linda; Larsen, Linda J.

1969-01-01

Although the Neisser—Wechsberg phenomenon resembles the inhibition of agglutination systems by excess antibody, the experimental results have indicated that with heat-inactivated antiserum the phenomenon results both from an excess of specific antibody and of non-specific anti-complementary activity. Complement (C) has been shown to be fixed in the presence of the excess antiserum which inhibits the bactericidal reaction. The inhibition was overcome by an excess of the third complement component factors indicating that the excess of antiserum interfered with the activation or function of the components acting at one of the late steps in the reaction sequence. The prozone phenomenon was relatively slight when unheated antiserum was used or when sensitized organisms were washed to remove serum substances unrelated to antibody. Non-specific anti-complementary activity, therefore, is a major contributor to the prozone phenomenon. Both IgM and IgG fractions of rabbit antisera elicited a prozone although the former had relatively greater bactericidal than inhibitory activity. PMID:5352361

Cpa, the outer membrane protease of Cronobacter sakazakii, activates plasminogen and mediates resistance to serum bactericidal activity.

PubMed

Franco, A A; Kothary, M H; Gopinath, G; Jarvis, K G; Grim, C J; Hu, L; Datta, A R; McCardell, B A; Tall, B D

2011-04-01

Cronobacter spp. are emerging neonatal pathogens in humans, associated with outbreaks of meningitis and sepsis. To cause disease, they must survive in blood and invade the central nervous system by penetrating the blood-brain barrier. C. sakazakii BAA-894 possesses an ~131-kb plasmid (pESA3) that encodes an outer membrane protease (Cpa) that has significant identity to proteins that belong to the Pla subfamily of omptins. Members of this subfamily of proteins degrade a number of serum proteins, including circulating complement, providing protection from the complement-dependent serum killing. Moreover, proteins of the Pla subfamily can cause uncontrolled plasmin activity by converting plasminogen to plasmin and inactivating the plasmin inhibitor α2-antiplasmin (α2-AP). These reactions enhance the spread and invasion of bacteria in the host. In this study, we found that an isogenic cpa mutant showed reduced resistance to serum in comparison to its parent C. sakazakii BAA-894 strain. Overexpression of Cpa in C. sakazakii or Escherichia coli DH5α showed that Cpa proteolytically cleaved complement components C3, C3a, and C4b. Furthermore, a strain of C. sakazakii overexpressing Cpa caused a rapid activation of plasminogen and inactivation of α2-AP. These results strongly suggest that Cpa may be an important virulence factor involved in serum resistance, as well as in the spread and invasion of C. sakazakii.

Mechanisms of action of escapin, a bactericidal agent in the ink secretion of the sea hare Aplysia californica: rapid and long-lasting DNA condensation and involvement of the OxyR-regulated oxidative stress pathway.

PubMed

Ko, Ko-Chun; Tai, Phang C; Derby, Charles D

2012-04-01

The marine snail Aplysia californica produces escapin, an L-amino acid oxidase, in its defensive ink. Escapin uses L-lysine to produce diverse products called escapin intermediate products of L-lysine (EIP-K), including α-amino-ε-caproic acid, Δ¹-piperidine-2-carboxylic acid, and Δ²-piperidine-2-carboxylic acid. EIP-K and H₂O₂ together, but neither alone, is a powerful bactericide. Here, we report bactericidal mechanisms of escapin products on Escherichia coli. We show that EIP-K and H₂O₂ together cause rapid and long-lasting DNA condensation: 2-min treatment causes significant DNA condensation and killing, and 10-min treatment causes maximal effect, lasting at least 70 h. We isolated two mutants resistant to EIP-K plus H₂O₂, both having a single missense mutation in the oxidation regulatory gene, oxyR. A complementation assay showed that the mutated gene, oxyR(A233V), renders resistance to EIP-K plus H₂O₂, and a gene dosage effect leads to reduction of resistance for strains carrying wild-type oxyR. Temperature stress with EIP-K does not produce the bactericidal effect, suggesting the effect is due to a specific response to oxidative stress. The null mutant for any single DNA-binding protein--Dps, H-NS, Hup, Him, or MukB--was not resistant to EIP-K plus H₂O₂, suggesting that no single DNA-binding protein is necessary to mediate this bactericidal effect, but allowing for the possibility that EIP-K plus H₂O₂ could function through a combination of DNA-binding proteins. The bactericidal effect of EIP-K plus H₂O₂ was eliminated by the ferrous ion chelator 1,10-phenanthroline, and it was reduced by the hydroxyl radical scavenger thiourea, suggesting hydroxyl radicals mediate the effects of EIP-K plus H₂O₂.

Structure-function characterization and optimization of a plant-derived antibacterial peptide.

PubMed

Suarez, Mougli; Haenni, Marisa; Canarelli, Stéphane; Fisch, Florian; Chodanowski, Pierre; Servis, Catherine; Michielin, Olivier; Freitag, Ruth; Moreillon, Philippe; Mermod, Nicolas

2005-09-01

Crushed seeds of the Moringa oleifera tree have been used traditionally as natural flocculants to clarify drinking water. We previously showed that one of the seed peptides mediates both the sedimentation of suspended particles such as bacterial cells and a direct bactericidal activity, raising the possibility that the two activities might be related. In this study, the conformational modeling of the peptide was coupled to a functional analysis of synthetic derivatives. This indicated that partly overlapping structural determinants mediate the sedimentation and antibacterial activities. Sedimentation requires a positively charged, glutamine-rich portion of the peptide that aggregates bacterial cells. The bactericidal activity was localized to a sequence prone to form a helix-loop-helix structural motif. Amino acid substitution showed that the bactericidal activity requires hydrophobic proline residues within the protruding loop. Vital dye staining indicated that treatment with peptides containing this motif results in bacterial membrane damage. Assembly of multiple copies of this structural motif into a branched peptide enhanced antibacterial activity, since low concentrations effectively kill bacteria such as Pseudomonas aeruginosa and Streptococcus pyogenes without displaying a toxic effect on human red blood cells. This study thus identifies a synthetic peptide with potent antibacterial activity against specific human pathogens. It also suggests partly distinct molecular mechanisms for each activity. Sedimentation may result from coupled flocculation and coagulation effects, while the bactericidal activity would require bacterial membrane destabilization by a hydrophobic loop.

Structure-Function Characterization and Optimization of a Plant-Derived Antibacterial Peptide

PubMed Central

Suarez, Mougli; Haenni, Marisa; Canarelli, Stéphane; Fisch, Florian; Chodanowski, Pierre; Servis, Catherine; Michielin, Olivier; Freitag, Ruth; Moreillon, Philippe; Mermod, Nicolas

2005-01-01

Crushed seeds of the Moringa oleifera tree have been used traditionally as natural flocculants to clarify drinking water. We previously showed that one of the seed peptides mediates both the sedimentation of suspended particles such as bacterial cells and a direct bactericidal activity, raising the possibility that the two activities might be related. In this study, the conformational modeling of the peptide was coupled to a functional analysis of synthetic derivatives. This indicated that partly overlapping structural determinants mediate the sedimentation and antibacterial activities. Sedimentation requires a positively charged, glutamine-rich portion of the peptide that aggregates bacterial cells. The bactericidal activity was localized to a sequence prone to form a helix-loop-helix structural motif. Amino acid substitution showed that the bactericidal activity requires hydrophobic proline residues within the protruding loop. Vital dye staining indicated that treatment with peptides containing this motif results in bacterial membrane damage. Assembly of multiple copies of this structural motif into a branched peptide enhanced antibacterial activity, since low concentrations effectively kill bacteria such as Pseudomonas aeruginosa and Streptococcus pyogenes without displaying a toxic effect on human red blood cells. This study thus identifies a synthetic peptide with potent antibacterial activity against specific human pathogens. It also suggests partly distinct molecular mechanisms for each activity. Sedimentation may result from coupled flocculation and coagulation effects, while the bactericidal activity would require bacterial membrane destabilization by a hydrophobic loop. PMID:16127062

Cpa, the Outer Membrane Protease of Cronobacter sakazakii, Activates Plasminogen and Mediates Resistance to Serum Bactericidal Activity▿

PubMed Central

Franco, A. A.; Kothary, M. H.; Gopinath, G.; Jarvis, K. G.; Grim, C. J.; Hu, L.; Datta, A. R.; McCardell, B. A.; Tall, B. D.

2011-01-01

Cronobacter spp. are emerging neonatal pathogens in humans, associated with outbreaks of meningitis and sepsis. To cause disease, they must survive in blood and invade the central nervous system by penetrating the blood-brain barrier. C. sakazakii BAA-894 possesses an ∼131-kb plasmid (pESA3) that encodes an outer membrane protease (Cpa) that has significant identity to proteins that belong to the Pla subfamily of omptins. Members of this subfamily of proteins degrade a number of serum proteins, including circulating complement, providing protection from the complement-dependent serum killing. Moreover, proteins of the Pla subfamily can cause uncontrolled plasmin activity by converting plasminogen to plasmin and inactivating the plasmin inhibitor α2-antiplasmin (α2-AP). These reactions enhance the spread and invasion of bacteria in the host. In this study, we found that an isogenic cpa mutant showed reduced resistance to serum in comparison to its parent C. sakazakii BAA-894 strain. Overexpression of Cpa in C. sakazakii or Escherichia coli DH5α showed that Cpa proteolytically cleaved complement components C3, C3a, and C4b. Furthermore, a strain of C. sakazakii overexpressing Cpa caused a rapid activation of plasminogen and inactivation of α2-AP. These results strongly suggest that Cpa may be an important virulence factor involved in serum resistance, as well as in the spread and invasion of C. sakazakii. PMID:21245266

An ecoimmunological approach to study evolutionary and ancient links between coagulation, complement and Innate immunity

PubMed Central

Kasetty, Gopinath; Alyafei, Saud; Smeds, Emanuel; Salo-Ahen, Outi M. H.; Hansson, Stefan R.; Egesten, Arne; Herwald, Heiko

2018-01-01

ABSTRACT Coagulation, complement, and innate immunity are tightly interwoven and form an alliance that can be traced back to early eukaryotic evolution. Here we employed an ecoimmunological approach using Tissue Factor Pathway Inhibitor (TFPI)-1-derived peptides from the different classes of vertebrates (i.e. fish, reptile, bird, and mammals) and tested whether they can boost killing of various human bacterial pathogens in plasma. We found signs of species-specific conservation and diversification during evolution in these peptides that significantly impact their antibacterial activity. Though all peptides tested executed bactericidal activity in mammalian plasma (with the exception of rodents), no killing was observed in plasma from birds, reptiles, and fish, pointing to a crucial role for the classical pathway of the complement system. We also observed an interference of these peptides with the human intrinsic pathway of coagulation though, unlike complement activation, this mechanism appears not to be evolutionary conserved. PMID:29473457

Mechanisms of Action of Escapin, a Bactericidal Agent in the Ink Secretion of the Sea Hare Aplysia californica: Rapid and Long-Lasting DNA Condensation and Involvement of the OxyR-Regulated Oxidative Stress Pathway

PubMed Central

Ko, Ko-Chun; Tai, Phang C.

2012-01-01

The marine snail Aplysia californica produces escapin, an l-amino acid oxidase, in its defensive ink. Escapin uses l-lysine to produce diverse products called escapin intermediate products of l-lysine (EIP-K), including α-amino-ε-caproic acid, Δ1-piperidine-2-carboxylic acid, and Δ2-piperidine-2-carboxylic acid. EIP-K and H2O2 together, but neither alone, is a powerful bactericide. Here, we report bactericidal mechanisms of escapin products on Escherichia coli. We show that EIP-K and H2O2 together cause rapid and long-lasting DNA condensation: 2-min treatment causes significant DNA condensation and killing, and 10-min treatment causes maximal effect, lasting at least 70 h. We isolated two mutants resistant to EIP-K plus H2O2, both having a single missense mutation in the oxidation regulatory gene, oxyR. A complementation assay showed that the mutated gene, oxyR(A233V), renders resistance to EIP-K plus H2O2, and a gene dosage effect leads to reduction of resistance for strains carrying wild-type oxyR. Temperature stress with EIP-K does not produce the bactericidal effect, suggesting the effect is due to a specific response to oxidative stress. The null mutant for any single DNA-binding protein—Dps, H-NS, Hup, Him, or MukB—was not resistant to EIP-K plus H2O2, suggesting that no single DNA-binding protein is necessary to mediate this bactericidal effect, but allowing for the possibility that EIP-K plus H2O2 could function through a combination of DNA-binding proteins. The bactericidal effect of EIP-K plus H2O2 was eliminated by the ferrous ion chelator 1,10-phenanthroline, and it was reduced by the hydroxyl radical scavenger thiourea, suggesting hydroxyl radicals mediate the effects of EIP-K plus H2O2. PMID:22232273

Ex vivo model of meningococcal bacteremia using human blood for measuring vaccine-induced serum passive protective activity.

PubMed

Plested, Joyce S; Welsch, Jo Anne; Granoff, Dan M

2009-06-01

The binding of complement factor H (fH) to meningococci was recently found to be specific for human fH. Therefore, passive protective antibody activity measured in animal models of meningococcal bacteremia may overestimate protection in humans, since in the absence of bound fH, complement activation is not downregulated. We developed an ex vivo model of meningococcal bacteremia using nonimmune human blood to measure the passive protective activity of stored sera from 36 adults who had been immunized with an investigational meningococcal multicomponent recombinant protein vaccine. Before immunization, human complement-mediated serum bactericidal activity (SBA) titers of > or = 1:4 against group B strains H44/76, NZ98/254, and S3032 were present in 19, 11, and 8% of subjects, respectively; these proportions increased to 97, 22, and 36%, respectively, 1 month after dose 3 (P < 0.01 for H44/76 and S3032). Against the two SBA-resistant strains, NZ98/254 and S3032, passive protective titers of > or = 1:4 were present in 11 and 42% of sera before immunization, respectively, and these proportions increased to 61 and 94% after immunization (P < 0.001 for each strain). Most of the sera with SBA titers of <1:4 and passive protective activity showed a level of killing in the whole-blood assay (>1 to 2 log(10) decreases in CFU/ml during a 90-min incubation) similar to that of sera with SBA titers of > or = 1:4. In conclusion, passive protective activity was 2.6- to 2.8-fold more frequent than SBA after immunization. The ability of SBA-negative sera to kill Neisseria meningitidis in human blood where fH is bound to the bacteria provides further evidence that SBA titers of > or = 1:4 measured with human complement may underestimate meningococcal immunity.

NETosing Neutrophils Activate Complement Both on Their Own NETs and Bacteria via Alternative and Non-alternative Pathways

PubMed Central

Yuen, Joshua; Pluthero, Fred G.; Douda, David N.; Riedl, Magdalena; Cherry, Ahmed; Ulanova, Marina; Kahr, Walter H. A.; Palaniyar, Nades; Licht, Christoph

2016-01-01

Neutrophils deposit antimicrobial proteins, such as myeloperoxidase and proteases on chromatin, which they release as neutrophil extracellular traps (NETs). Neutrophils also carry key components of the complement alternative pathway (AP) such as properdin or complement factor P (CFP), complement factor B (CFB), and C3. However, the contribution of these complement components and complement activation during NET formation in the presence and absence of bacteria is poorly understood. We studied complement activation on NETs and a Gram-negative opportunistic bacterial pathogen Pseudomonas aeruginosa (PA01, PAKwt, and PAKgfp). Here, we show that anaphylatoxin C5a, formyl-methionyl-leucyl-phenylalanine (fMLP) and phorbol myristate acetate (PMA), which activates NADPH oxidase, induce the release of CFP, CFB, and C3 from neutrophils. In response to PMA or P. aeruginosa, neutrophils secrete CFP, deposit it on NETs and bacteria, and induce the formation of terminal complement complexes (C5b–9). A blocking anti-CFP antibody inhibited AP-mediated but not non-AP-mediated complement activation on NETs and P. aeruginosa. Therefore, NET-mediated complement activation occurs via both AP- and non AP-based mechanisms, and AP-mediated complement activation during NETosis is dependent on CFP. These findings suggest that neutrophils could use their “AP tool kit” to readily activate complement on NETs and Gram-negative bacteria, such as P. aeruginosa, whereas additional components present in the serum help to fix non-AP-mediated complement both on NETs and bacteria. This unique mechanism may play important roles in host defense and help to explain specific roles of complement activation in NET-related diseases. PMID:27148258

Human Lipooligosaccharide IGG That Prevents Endemic Meningococcal Disease Recognizes an Internal Lacto-N-neotetraose Structure*

PubMed Central

Cheng, Hui; Yang, Zhijie; Estabrook, Michele M.; John, Constance M.; Jarvis, Gary A.; McLaughlin, Stephanie; Griffiss, J. McLeod

2011-01-01

Antibodies that initiate complement-mediated killing of Neisseria meningitidis as they enter the bloodstream from the oropharynx protect against disseminated disease. Human IgGs that bind the neisserial L7 lipooligosaccharide (LOS) are bactericidal for L3,7 and L2,4 meningococci in the presence of human complement. These strains share a lacto-N-neotetraose (nLc4) LOS α chain. We used a set of mutants that have successive saccharide deletions from the nLc4 α chain to characterize further the binding and bactericidal activity of nLc4 LOS IgG. We found that the nLc4 α chain conforms at least four different antigens. We separately purified IgG that required the nLc4 (non-reducing) terminal galactose (Gal) for binding and IgG that bound the truncated nLc3 α chain that lacks this Gal residue. IgG that bound the internal nLc3 α chain killed both L3,7 and L2,4 strains, whereas IgG that required the nLc4 terminal Gal residue for binding killed L2,4 stains but not L3,7 strains. These results show that the diversity of LOS antibodies in human serum is as much a function of the conformation of multiple antigens by a single glycoform as of the production of multiple glycoforms. Differences in sensitivity to killing by human nLc4 LOS IgG may account for the fact that fully two-thirds of endemic group B meningococcal disease in infants and children is caused by L3,7 strains, but only 20% is caused by L2,4 stains. PMID:22027827

Peroxidasin contributes to lung host defense by direct binding and killing of gram-negative bacteria.

PubMed

Shi, Ruizheng; Cao, Zehong; Li, Hong; Graw, Jochen; Zhang, Guogang; Thannickal, Victor J; Cheng, Guangjie

2018-05-01

Innate immune recognition is classically mediated by the interaction of host pattern-recognition receptors and pathogen-associated molecular patterns; this triggers a series of downstream signaling events that facilitate killing and elimination of invading pathogens. In this report, we provide the first evidence that peroxidasin (PXDN; also known as vascular peroxidase-1) directly binds to gram-negative bacteria and mediates bactericidal activity, thus, contributing to lung host defense. PXDN contains five leucine-rich repeats and four immunoglobulin domains, which allows for its interaction with lipopolysaccharide, a membrane component of gram-negative bacteria. Bactericidal activity of PXDN is mediated via its capacity to generate hypohalous acids. Deficiency of PXDN results in a failure to eradicate Pseudomonas aeruginosa and increased mortality in a murine model of Pseudomonas lung infection. These observations indicate that PXDN mediates previously unrecognized host defense functions against gram-negative bacterial pathogens.

In vitro and in vivo activities of E-101 solution against Acinetobacter baumannii isolates from U.S. military personnel.

PubMed

Denys, G A; Davis, J C; O'Hanley, P D; Stephens, J T

2011-07-01

We evaluated the in vitro and in vivo activity of a novel topical myeloperoxidase-mediated antimicrobial, E-101 solution, against 5 multidrug-resistant Acinetobacter baumannii isolates recovered from wounded American soldiers. Time-kill studies demonstrated rapid bactericidal activity against all A. baumannii strains tested in the presence of 3% blood. The in vitro bactericidal activity of E-101 solution against A. baumannii strains was confirmed in a full-thickness excision rat model. Additional in vivo studies appear warranted.

Bactericidal catechins damage the lipid bilayer.

PubMed

Ikigai, H; Nakae, T; Hara, Y; Shimamura, T

1993-04-08

The mode of antibacterial action of, the green tea (Camellia sinensis) extracts, (-)-epigallocatechin gallate (EGCg) and (-)-epicatechin (EC) was investigated. Strong bactericidal EGCg caused leakage of 5,6-carboxyfluorescein from phosphatidylcholine liposomes (PC), but EC with very weak bactericidal activity caused little damage to the membrane. Phosphatidylserine and dicetyl phosphate partially protected the membrane from EGCg-mediated damage when reconstituted into the liposome membrane with PC. EGCg, but not EC, caused strong aggregation and NPN-fluorescence quenching of PC-liposomes and these actions were markedly lowered in the presence of negatively charged lipids. These results show that bactericidal catechins primarily act on and damage bacterial membranes. The observation that Gram-negative bacteria are more resistant to bactericidal catechins than Gram-positive bacteria can be explained to some extent by the presence of negatively charged lipopolysaccharide.

Minimum structural requirements for cell membrane leakage-mediated anti-MRSA activity of macrocyclic bis(bibenzyl)s.

PubMed

Fujii, Kana; Morita, Daichi; Onoda, Kenji; Kuroda, Teruo; Miyachi, Hiroyuki

2016-05-01

Macrocyclic bis(bibenzyl)-type phenolic natural products, found exclusively in bryophytes, exhibit potent antibacterial activity towards methicillin-resistant Staphylococcus aureus (anti-MRSA activity). Here, in order to identify the minimum essential structure for cell membrane leakage-mediated anti-MRSA activity of these compounds, we synthesized acyclic fragment structures and evaluated their anti-MRSA activity. The activities of all of the acyclic fragments tested exhibited similar characteristics to those of the macrocycles, i.e., anti-MRSA bactericidal activity, an enhancing effect on influx and efflux of ethidium bromide (EtBr: fluorescent DNA-binder) in Staphylococcus aureus cells, and bactericidal activity towards a Staphylococcus aureus strain resistant to 2-phenoxyphenol (4). The latter result suggests that they have a different mechanism of action from 4, which is a FabI inhibitor previously proposed to be the minimum active fragment of riccardin-type macrocycles. Thus, cyclic structure is not a necessary condition for cell membrane leakage-mediated anti-MRSA activity of macrocyclic bis(bibenzyl)s. Copyright © 2016 Elsevier Ltd. All rights reserved.

Logarithmic phase Escherichia coli K1 efficiently avoids serum killing by promoting C4bp-mediated C3b and C4b degradation

PubMed Central

Wooster, David G; Maruvada, Ravi; Blom, Anna M; Prasadarao, Nemani V

2006-01-01

Meningitis caused by Escherichia coli K1 is a serious illness in neonates with neurological sequelae in up to 50% of survivors. A high degree of bacteremia is required for E. coli K1 to cross the blood–brain barrier, which suggests that the bacterium must evade the host defence mechanisms and survive in the bloodstream. We previously showed that outer membrane protein A (OmpA) of E. coli binds C4b-binding protein (C4bp), an inhibitor of complement activation via the classical pathway. Nevertheless, the exact mechanism by which E. coli K1 survives in serum remains elusive. Here, we demonstrate that log phase (LP) OmpA+E. coli K1 avoids serum bactericidal activity more effectively than postexponential phase bacteria. OmpA–E. coli cannot survive in serum grown to either phase. The increased serum resistance of LP OmpA+E. coli is the result of increased binding of C4bp, with a concomitant decrease in the deposition of C3b and the downstream complement proteins responsible for the formation of the membrane attack complex. C4bp bound to E. coli K1 acts as a cofactor to factor I in the cleavage of both C3b and C4b, which shuts down the ensuing complement cascade. Accordingly, a peptide corresponding to the complement control protein domain 3 of C4bp sequence, was able to compete with C4bp binding to OmpA and cause increased deposition of C3b. Thus, binding of C4bp appears to be responsible for survival of E. coli K1 in human serum. PMID:16556262

Effects of dietary vitamin C on the immune function of shrimps, Penaeus chinensis

NASA Astrophysics Data System (ADS)

Wang, Weiqing; Li, Aijie; Cheung, Siugin

2002-04-01

Prepared in this experiment were six groups of diets, i.e. VC0, VC1, VC2, VC3, VC4 and VC5 with the contents of vitamin C (VCmg(100 g)-1 diet) of 0, 100, 200, 400, 800, and 1200 respectively. It was found that vitamin C increased the concentrations of immunoglobulin-like (IgG-like, IgA-like and IgM-like) substances in the serum of Penaeus chinensis after a feeding period of 3 weeks. The differences among groups were significant (P<0.01), but there was no difference in the contents of complement3-like and complement4-like substances in the serum (P>0.05). Phenoloxidase (PO) activity in the serum of VC3 group shrimps was higher than that of VC0 and other groups, but no significant difference was observed between VC0 group and other groups. Furthermore, bactericidal activity of the serum to Vibrio parahaemolyticus in shrimps fed with the VC1 diet was higher than that in the other groups (P<0.01), while no difference was demonstrated among all groups for the bactericidal activity to Vibrio alginolyticus (P>0.05). It is, therefore, suggested that vitamin C (100 400 mg(100 g)-1 diets) could be used as an immunostimulant of P. chinensis.

Effectiveness analyses may underestimate protection of infants after group C meningococcal immunization.

PubMed

Vu, David M; Kelly, Dominic; Heath, Paul T; McCarthy, Noel D; Pollard, Andrew J; Granoff, Dan M

2006-07-15

Group C meningococcal conjugate-vaccine effectiveness in the United Kingdom declines from ~90% in the first year to 0% between 1 and 4 years after immunization in infants immunized at 2, 3, and 4 months of age and to 61% in toddlers given a single dose. Confidence intervals are wide, and the extent of protection is uncertain. Serum samples were obtained from children 3-5 years of age who were participants in a preschool booster-vaccine trial. Serum bactericidal activity was measured with human complement. Group C anticapsular antibody concentrations were measured by a radioantigen binding assay. Passive protection was analyzed in an infant rat bacteremia model. Serum samples from UK children who had been immunized 2-3 years earlier as infants or toddlers had higher levels of radioantigen binding, bactericidal activity, and passive protection than did historical control serum samples from unimmunized children (P<.05). A higher proportion of children immunized as infants had serum bactericidal activity titers > or =1 : 4 (considered to be protective) than those immunized as toddlers (61% vs. 24%; P<.01), but there were no significant differences in the proportion of serum samples conferring passive protection (50% and 41%, respectively; P=.4). We found no evidence of lower immunity in children immunized as infants than as toddlers. On the basis of serum bactericidal activity and/or passive protection, 40%-50% of both age groups are protected at 2-3 years after immunization, which was significantly greater than in unimmunized historical controls (<5%).

Human antibodies fix complement to inhibit Plasmodium falciparum invasion of erythrocytes and are associated with protection against malaria.

PubMed

Boyle, Michelle J; Reiling, Linda; Feng, Gaoqian; Langer, Christine; Osier, Faith H; Aspeling-Jones, Harvey; Cheng, Yik Sheng; Stubbs, Janine; Tetteh, Kevin K A; Conway, David J; McCarthy, James S; Muller, Ivo; Marsh, Kevin; Anders, Robin F; Beeson, James G

2015-03-17

Antibodies play major roles in immunity to malaria; however, a limited understanding of mechanisms mediating protection is a major barrier to vaccine development. We have demonstrated that acquired human anti-malarial antibodies promote complement deposition on the merozoite to mediate inhibition of erythrocyte invasion through C1q fixation and activation of the classical complement pathway. Antibody-mediated complement-dependent (Ab-C') inhibition was the predominant invasion-inhibitory activity of human antibodies; most antibodies were non-inhibitory without complement. Inhibitory activity was mediated predominately via C1q fixation, and merozoite surface proteins 1 and 2 were identified as major targets. Complement fixation by antibodies was very strongly associated with protection from both clinical malaria and high-density parasitemia in a prospective longitudinal study of children. Ab-C' inhibitory activity could be induced by human immunization with a candidate merozoite surface-protein vaccine. Our findings demonstrate that human anti-malarial antibodies have evolved to function by fixing complement for potent invasion-inhibitory activity and protective immunity. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

[Effect of repeated cycles of tetraolean and oleandomycin administration on the indices of nonspecific resistence of the body in experimental staphylococcal sepsis].

PubMed

Plaksin, A I; Plaksina, A N

1975-02-01

The effect of repeated cycles of tetracycline and oleandomycin administration on the complement titer, content of lysozyme and bactericidal properties of the serum in rabbits with experimental staphylococcal sepsis was studied. It was shown that the septic process induced by intravenous inoculation of staphylococci was accompanied by stimulation of the host nonspecific resistance. However, repeated inoculations of the animals resulted in exhaustion of the host protective forces and decreased non-specific resistance. The use of tetracycline in experimental staphylococcal sepsis was accompanied by an increase in the complement titer, lysozyme content and bactericidal properties of the serum after both the 1st and 2nd cycles of the drug administration. The use of oleaudomycin induced an increase in the contents of the complement, lysozyme and bactericidal properties of the serum at the background of staphylococcal sepsis only after the 1st cycle. The repeated cycle of oleandomycin administration was accompanied by a decrease in the above indices. Such conditions should be taken into account in choosing the antibiotic for treatment of septic cases especially when repeated cycles of the drug administration are used.

Antibody persistence for up to 5 years after a fourth dose of Haemophilus influenzae type b and Neisseria meningitidis serogroups C and Y-tetanus toxoid conjugate vaccine (HibMenCY-TT) given at 12-15 months of age.

PubMed

Marshall, Gary S; Blatter, Mark; Marchant, Colin; Aris, Emmanuel; Mesaros, Narcisa; Miller, Jacqueline M

2013-06-01

A 4-dose series of recently licensed Haemophilus influenzae type b-meningococcal serogroups C and Y-tetanus toxoid conjugate vaccine (HibMenCY-TT) was immunogenic with a clinically acceptable safety profile in infants, with antibodies persisting in most participants for 1 year following dose 4. This study assessed antibody persistence up to 5 years after vaccination. Participants had received HibMenCY-TT or Hib-TT at 2, 4 and 6 months of age. At age 12-15 months, HibMenCY-TT vaccinees received a fourth HibMenCY-TT dose (HibMenCY x 4 group), whereas those who received Hib-TT received a fourth dose of either Hib-TT (Hib) or HibMenCY-TT (HibMenCY x 1). Blood samples were collected 1 month and 1, 3 and 5 years after the last dose for measurement of antipolyribosylribitol phosphate (the Hib capsular polysaccharide) antibodies and serum bactericidal activity (human complement source) against meningococcal serogroups C and Y. Five years after the fourth dose, the percentages of children with antipolyribosylribitol phosphate ≥0.15 μg/mL in HibMenCY x 4, HibMenCY x 1 and Hib groups were 98.8% (95% confidence interval: 93.5%-100%), 97.3% (85.8%-99.9%) and 92.3% (79.1%-98.4%), respectively. The percentages with human complement serum bactericidal activity ≥1:8 for meningococcal serogroup C were 82.9% (72.5%-90.6%), 73.5% (55.6%-87.1%) and 21.1% (9.6%-37.3%), respectively. The percentages with human complement serum bactericidal activity ≥1:8 for serogroup Y were 69.5% (58.4%-79.2%), 54.3% (36.6%-71.2%) and 18.4% (7.7%-34.3%), respectively. HibMenCY-TT given as a 4-dose series or as a single dose at 12-15 months of age induced immune responses for all 3 antigens that lasted for up to 5 years after vaccination in more than half of recipients.

Natural Product Anacardic Acid from Cashew Nut Shells Stimulates Neutrophil Extracellular Trap Production and Bactericidal Activity.

PubMed

Hollands, Andrew; Corriden, Ross; Gysler, Gabriela; Dahesh, Samira; Olson, Joshua; Raza Ali, Syed; Kunkel, Maya T; Lin, Ann E; Forli, Stefano; Newton, Alexandra C; Kumar, Geetha B; Nair, Bipin G; Perry, J Jefferson P; Nizet, Victor

2016-07-01

Emerging antibiotic resistance among pathogenic bacteria is an issue of great clinical importance, and new approaches to therapy are urgently needed. Anacardic acid, the primary active component of cashew nut shell extract, is a natural product used in the treatment of a variety of medical conditions, including infectious abscesses. Here, we investigate the effects of this natural product on the function of human neutrophils. We find that anacardic acid stimulates the production of reactive oxygen species and neutrophil extracellular traps, two mechanisms utilized by neutrophils to kill invading bacteria. Molecular modeling and pharmacological inhibitor studies suggest anacardic acid stimulation of neutrophils occurs in a PI3K-dependent manner through activation of surface-expressed G protein-coupled sphingosine-1-phosphate receptors. Neutrophil extracellular traps produced in response to anacardic acid are bactericidal and complement select direct antimicrobial activities of the compound. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Guilty as charged: all available evidence implicates complement's role in fetal demise.

PubMed

Girardi, Guillermina

2008-03-01

Appropriate complement inhibition is an absolute requirement for normal pregancy. Uncontrolled complement activation in the maternal-fetal interface leads to fetal death. Here we show that complement activation is a crucial and early mediator of pregnancy loss in two different mouse models of pregnancy loss. Using a mouse model of fetal loss and growth restriction (IUGR) induced by antiphospholipid antibodies (aPL), we examined the role of complement activation in fetal loss and IUGR. We found that C5a-C5aR interaction and neutrophils are key mediators of fetal injury. Treatment with heparin, the standard therapy for pregnant patients with aPL, prevents complement activation and protects mice from pregnancy complications induced by aPL, and anticoagulants that do not inhibit complement do not protect pregnancies. In an antibody-independent mouse model of spontaneous miscarriage and IUGR (CBA/JxDBA/2) we also identified C5a as an essential mediator. Complement activation caused dysregulation of the angiogenic factors required for normal placental development. In CBA/JxDBA/2 mice, we observed inflammatory infiltrates in placentas, functional deficiency of free vascular endothelial growth factor (VEGF), elevated levels of soluble VEGF receptor-1 (sVEGFR-1, also known as sFlt-1; a potent anti-angiogenic molecule), and defective placental development. Inhibition of complement activation blocked the increase in sVEGFR-1 and rescued pregnancies. Our studies in antibody-dependent and antibody-independent models of pregnancy complications identified complement activation as the key mediator of damage and will allow development of new interventions to prevent pregnancy loss and IUGR.

Evasion of Complement-Mediated Lysis and Complement C3 Deposition Are Regulated by Francisella tularensis Lipopolysaccharide O Antigen1

PubMed Central

Clay, Corey D.; Soni, Shilpa; Gunn, John S.; Schlesinger, Larry S.

2009-01-01

The bacterium Francisella tularensis (Ft) is a potential weapon of bioterrorism when aerosolized. Macrophage infection is necessary for disease progression and efficient phagocytosis by human macrophages requires serum opsonization by complement. Microbial complement activation leads to surface deposition of a highly regulated protein complex resulting in opsonization or membrane lysis. The nature of complement component C3 deposition, i.e., C3b (opsonization and lysis) or C3bi (opsonization only) fragment deposition, is central to the outcome of activation. In this study, we examine the mechanisms of Ft resistance to complement-mediated lysis, C3 component deposition on the Ft surface, and complement activation. Upon incubation in fresh nonimmune human serum, Schu S4 (Ft subsp. tularensis), Fn (Ft subsp. novicida), and LVS (Ft subsp. holarctica live vaccine strain) were resistant to complement-mediated lysis, but LVSG and LVSR (LVS strains altered in surface carbohydrate structures) were susceptible. C3 deposition, however, occurred on all strains. Complement-susceptible strains had markedly increased C3 fragment deposition, including the persistent presence of C3b compared with C3bi, which indicates that C3b inactivation results in survival of complement-resistant strains. C1q, an essential component of the classical activation pathway, was necessary for lysis of complement-susceptible strains and optimal C3 deposition on all strains. Finally, use of Francisella LPS mutants confirmed O Ag as a major regulator of complement resistance. These data provide evidence that pathogenic Francisella activate complement, but are resistant to complement-mediated lysis in part due to limited C3 deposition, rapid conversion of surface-bound C3b to C3bi, and the presence of LPS O Ag. PMID:18832715

CsMAP34, a teleost MAP with dual role: A promoter of MASP-assisted complement activation and a regulator of immune cell activity.

PubMed

Li, Mo-Fei; Li, Jun; Sun, Li

2016-12-23

In teleost fish, the immune functions of mannan-binding lectin (MBL) associated protein (MAP) and MBL associated serine protease (MASP) are scarcely investigated. In the present study, we examined the biological properties both MAP (CsMAP34) and MASP (CsMASP1) molecules from tongue sole (Cynoglossus semilaevis). We found that CsMAP34 and CsMASP1 expressions occurred in nine different tissues and were upregulated by bacterial challenge. CsMAP34 protein was detected in blood, especially during bacterial infection. Recombinant CsMAP34 (rCsMAP34) bound C. semilaevis MBL (rCsBML) when the latter was activated by bacteria, while recombinant CsMASP1 (rCsMASP1) bound activated rCsBML only in the presence of rCsMAP34. rCsMAP34 stimulated the hemolytic and bactericidal activities of serum complement, whereas anti-CsMAP34 antibody blocked complement activities. Knockdown of CsMASP1 in C. semilaevis resulted in significant inhibition of complement activities. Furthermore, rCsMAP34 interacted directly with peripheral blood leukocytes (PBL) and enhanced the respiratory burst, acid phosphatase activity, chemotactic activity, and gene expression of PBL. These results indicate for the first time that a teleost MAP acts one hand as a regulator that promotes the lectin pathway of complement activation via its ability to recruit MBL to MASP, and other hand as a modulator of immune cell activity.

Selenium-mediated protection in reversing the sensitivity of bacterium to the bactericidal antibiotics.

PubMed

Li, Zhonglei; Tan, Jun; Shao, Lei; Dong, Xiaojing; Ye, Richard D; Chen, Daijie

2017-05-01

Inducing production of damaging reactive oxygen species (ROS) is an important criterion to distinguish the bactericidal antibiotics from bacteriostatic antibiotics. Selenoenzymes were generally recognized to be a powerful antioxidant capable of scavenging free radicals, protecting the cells from the harmful effects of ROS. Therefore, the present study was carried out to investigate the selenium (Se)-mediated protection in reversing antibiotic sensitivity and the role of selenoenzymes in alleviating the negative effects of oxidative stress. The cellular antioxidant activity of Se-enriched bacteria was analyzed, as well as intracellular ROS production and elimination when Se-enriched bacteria in the presence of various antibiotics. Compared to complete inhibition of the parental strain by bactericidal antibiotics, it only exhibited slight and reversible inhibition of Se-enriched Escherichia coli ATCC25922 and Staphylococcus aureus ATCC25923 at the same conditions, which indicated that intracellular selenium provided substantial protection against antibiotics. ROS generation caused by bactericidal antibiotics was confirmed by fluorescence spectrophotometry using 2', 7'-dichloro- uorescein diacetate (DCFH-DA) as substrate. The time course experiments of pretreatment with selenium showed significant decrease of ROS level at 2h. In summary, the present study provides experimental evidence supporting selenoenzymes has good scavenging effect to ROS and can protect bacteria from oxidative stress injury induced by bactericidal antibiotics. Copyright © 2017 Elsevier GmbH. All rights reserved.

Complement in Non-Antibody-Mediated Kidney Diseases

PubMed Central

Angeletti, Andrea; Reyes-Bahamonde, Joselyn; Cravedi, Paolo; Campbell, Kirk N.

2017-01-01

The complement system is part of the innate immune response that plays important roles in protecting the host from foreign pathogens. The complement components and relative fragment deposition have long been recognized to be strongly involved also in the pathogenesis of autoantibody-related kidney glomerulopathies, leading to direct glomerular injury and recruitment of infiltrating inflammation pathways. More recently, unregulated complement activation has been shown to be associated with progression of non-antibody-mediated kidney diseases, including focal segmental glomerulosclerosis, C3 glomerular disease, thrombotic microangiopathies, or general fibrosis generation in progressive chronic kidney diseases. Some of the specific mechanisms associated with complement activation in these diseases were recently clarified, showing a dominant role of alternative activation pathway. Over the last decade, a growing number of anticomplement agents have been developed, and some of them are being approved for clinical use or already in use. Therefore, anticomplement therapies represent a realistic choice of therapeutic approaches for complement-related diseases. Herein, we review the complement system activation, regulatory mechanisms, their involvement in non-antibody-mediated glomerular diseases, and the recent advances in complement-targeting agents as potential therapeutic strategies. PMID:28748184

Methylfolate Trap Promotes Bacterial Thymineless Death by Sulfa Drugs

PubMed Central

Pham, Thanh H.; Jakubowski, Hieronim; Wolff, Kerstin A.; Ogwang, Sam; Timpona, Joseph L.; Gogula, Soumya; Jacobs, Michael R.; Ruetz, Markus; Kräutler, Bernhard; Jacobsen, Donald W.; Zhang, Guo-Fang; Nguyen, Liem

2016-01-01

The methylfolate trap, a metabolic blockage associated with anemia, neural tube defects, Alzheimer’s dementia, cardiovascular diseases, and cancer, was discovered in the 1960s, linking the metabolism of folate, vitamin B12, methionine and homocysteine. However, the existence or physiological significance of this phenomenon has been unknown in bacteria, which synthesize folate de novo. Here we identify the methylfolate trap as a novel determinant of the bacterial intrinsic death by sulfonamides, antibiotics that block de novo folate synthesis. Genetic mutagenesis, chemical complementation, and metabolomic profiling revealed trap-mediated metabolic imbalances, which induced thymineless death, a phenomenon in which rapidly growing cells succumb to thymine starvation. Restriction of B12 bioavailability, required for preventing trap formation, using an “antivitamin B12” molecule, sensitized intracellular bacteria to sulfonamides. Since boosting the bactericidal activity of sulfonamides through methylfolate trap induction can be achieved in Gram-negative bacteria and mycobacteria, it represents a novel strategy to render these pathogens more susceptible to existing sulfonamides. PMID:27760199

Symbiotic Bacteria Direct Expression of an Intestinal Bactericidal Lectin

PubMed Central

Cash, Heather L.; Whitham, Cecilia V.; Behrendt, Cassie L.; Hooper, Lora V.

2009-01-01

The mammalian intestine harbors complex societies of beneficial bacteria that are maintained in the lumen with minimal penetration of mucosal surfaces. Microbial colonization of germ-free mice triggers epithelial expression of RegIIIγ, a secreted C-type lectin. RegIIIγ binds intestinal bacteria but lacks the complement recruitment domains present in other microbe-binding mammalian C-type lectins. We show that RegIIIγ and its human counterpart, HIP/PAP, are directly antimicrobial proteins that bind their bacterial targets via interactions with peptidoglycan carbohydrate. We propose that these proteins represent an evolutionarily primitive form of lectin-mediated innate immunity, and that they reveal intestinal strategies for maintaining symbiotic host-microbial relationships. PMID:16931762

Human bactericidal/permeability-increasing protein and a recombinant NH2-terminal fragment cause killing of serum-resistant gram-negative bacteria in whole blood and inhibit tumor necrosis factor release induced by the bacteria.

PubMed Central

Weiss, J; Elsbach, P; Shu, C; Castillo, J; Grinna, L; Horwitz, A; Theofan, G

1992-01-01

The bactericidal/permeability-increasing protein (BPI) of neutrophils and BPI fragments neutralize the effects of isolated Gram-negative bacterial lipopolysaccharides both in vitro and in vivo. Since endotoxin most commonly enters the host as constituents of invading Gram-negative bacteria, we raised the question: Can BPI and its bioactive fragments also protect against whole bacteria? To determine whether the bactericidal and endotoxin-neutralizing activities of BPI/fragments are expressed when Gram-negative bacteria are introduced to the complex environment of whole blood we examined the effects of added BPI and proteolytically prepared and recombinant NH2-terminal fragments on: (a) the fate of serum-resistant encapsulated Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa that survive the antibacterial actions of whole blood and (b) the ability of these bacteria to trigger cytokine release. Added BPI in nanomolar concentrations killed each of three encapsulated strains of E. coli and in closely parallel fashion inhibited tumor necrosis factor (TNF) release. Holo-BPI and its NH2-terminal fragment were equipotent toward a rough LPS chemotype K1-encapsulated strain, but the fragment was substantially more potent than holo-BPI toward two encapsulated smooth LPS chemotype strains. TNF release induced by K. pneumoniae and P. aeruginosa was also inhibited by both holo-BPI and fragment but, at the protein concentrations tested, P. aeruginosa was killed only by the fragment and K. pneumoniae was not killed by either protein. The bactericidal action of BPI/fragment toward E. coli is inhibited by C7-depleted serum, but accelerated by normal serum, indicating that BPI, acting in synergy with late complement components, enhances extracellular killing of serum-resistant bacteria. Thus, BPI and an even more potent NH2-terminal fragment may protect against Gram-negative bacteria in the host by blocking bacterial proliferation as well as endotoxin-mediated effects, not only as components of the intracellular antibacterial arsenal of the neutrophil, but also as potentially therapeutic extracellular agents. PMID:1522221

Human bactericidal/permeability-increasing protein and a recombinant NH2-terminal fragment cause killing of serum-resistant gram-negative bacteria in whole blood and inhibit tumor necrosis factor release induced by the bacteria.

PubMed

Weiss, J; Elsbach, P; Shu, C; Castillo, J; Grinna, L; Horwitz, A; Theofan, G

1992-09-01

The bactericidal/permeability-increasing protein (BPI) of neutrophils and BPI fragments neutralize the effects of isolated Gram-negative bacterial lipopolysaccharides both in vitro and in vivo. Since endotoxin most commonly enters the host as constituents of invading Gram-negative bacteria, we raised the question: Can BPI and its bioactive fragments also protect against whole bacteria? To determine whether the bactericidal and endotoxin-neutralizing activities of BPI/fragments are expressed when Gram-negative bacteria are introduced to the complex environment of whole blood we examined the effects of added BPI and proteolytically prepared and recombinant NH2-terminal fragments on: (a) the fate of serum-resistant encapsulated Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa that survive the antibacterial actions of whole blood and (b) the ability of these bacteria to trigger cytokine release. Added BPI in nanomolar concentrations killed each of three encapsulated strains of E. coli and in closely parallel fashion inhibited tumor necrosis factor (TNF) release. Holo-BPI and its NH2-terminal fragment were equipotent toward a rough LPS chemotype K1-encapsulated strain, but the fragment was substantially more potent than holo-BPI toward two encapsulated smooth LPS chemotype strains. TNF release induced by K. pneumoniae and P. aeruginosa was also inhibited by both holo-BPI and fragment but, at the protein concentrations tested, P. aeruginosa was killed only by the fragment and K. pneumoniae was not killed by either protein. The bactericidal action of BPI/fragment toward E. coli is inhibited by C7-depleted serum, but accelerated by normal serum, indicating that BPI, acting in synergy with late complement components, enhances extracellular killing of serum-resistant bacteria. Thus, BPI and an even more potent NH2-terminal fragment may protect against Gram-negative bacteria in the host by blocking bacterial proliferation as well as endotoxin-mediated effects, not only as components of the intracellular antibacterial arsenal of the neutrophil, but also as potentially therapeutic extracellular agents.

Complement Activation in Inflammatory Skin Diseases

PubMed Central

Giang, Jenny; Seelen, Marc A. J.; van Doorn, Martijn B. A.; Rissmann, Robert; Prens, Errol P.; Damman, Jeffrey

2018-01-01

The complement system is a fundamental part of the innate immune system, playing a crucial role in host defense against various pathogens, such as bacteria, viruses, and fungi. Activation of complement results in production of several molecules mediating chemotaxis, opsonization, and mast cell degranulation, which can contribute to the elimination of pathogenic organisms and inflammation. Furthermore, the complement system also has regulating properties in inflammatory and immune responses. Complement activity in diseases is rather complex and may involve both aberrant expression of complement and genetic deficiencies of complement components or regulators. The skin represents an active immune organ with complex interactions between cellular components and various mediators. Complement involvement has been associated with several skin diseases, such as psoriasis, lupus erythematosus, cutaneous vasculitis, urticaria, and bullous dermatoses. Several triggers including auto-antibodies and micro-organisms can activate complement, while on the other hand complement deficiencies can contribute to impaired immune complex clearance, leading to disease. This review provides an overview of the role of complement in inflammatory skin diseases and discusses complement factors as potential new targets for therapeutic intervention. PMID:29713318

Fosfomycin enhances phagocyte-mediated killing of Staphylococcus aureus by extracellular traps and reactive oxygen species.

PubMed

Shen, Fengge; Tang, Xudong; Cheng, Wei; Wang, Yang; Wang, Chao; Shi, Xiaochen; An, Yanan; Zhang, Qiaoli; Liu, Mingyuan; Liu, Bo; Yu, Lu

2016-01-18

The successful treatment of bacterial infections is the achievement of a synergy between the host's immune defences and antibiotics. Here, we examined whether fosfomycin (FOM) could improve the bactericidal effect of phagocytes, and investigated the potential mechanisms. FOM enhanced the phagocytosis and extra- or intracellular killing of S. aureus by phagocytes. And FOM enhanced the extracellular killing of S. aureus in macrophage (MФ) and in neutrophils mediated by extracellular traps (ETs). ET production was related to NADPH oxidase-dependent reactive oxygen species (ROS). Additionally, FOM increased the intracellular killing of S. aureus in phagocytes, which was mediated by ROS through the oxidative burst process. Our results also showed that FOM alone induced S. aureus producing hydroxyl radicals in order to kill the bacterial cells in vitro. In a mouse peritonitis model, FOM treatment increased the bactericidal extra- and intracellular activity in vivo, and FOM strengthened ROS and ET production from peritoneal lavage fluid ex vivo. An IVIS imaging system assay further verified the observed in vivo bactericidal effect of the FOM treatment. This work may provide a deeper understanding of the role of the host's immune defences and antibiotic interactions in microbial infections.

Fosfomycin enhances phagocyte-mediated killing of Staphylococcus aureus by extracellular traps and reactive oxygen species

PubMed Central

Shen, Fengge; Tang, Xudong; Cheng, Wei; Wang, Yang; Wang, Chao; Shi, Xiaochen; An, Yanan; Zhang, Qiaoli; Liu, Mingyuan; Liu, Bo; Yu, Lu

2016-01-01

The successful treatment of bacterial infections is the achievement of a synergy between the host’s immune defences and antibiotics. Here, we examined whether fosfomycin (FOM) could improve the bactericidal effect of phagocytes, and investigated the potential mechanisms. FOM enhanced the phagocytosis and extra- or intracellular killing of S. aureus by phagocytes. And FOM enhanced the extracellular killing of S. aureus in macrophage (MФ) and in neutrophils mediated by extracellular traps (ETs). ET production was related to NADPH oxidase-dependent reactive oxygen species (ROS). Additionally, FOM increased the intracellular killing of S. aureus in phagocytes, which was mediated by ROS through the oxidative burst process. Our results also showed that FOM alone induced S. aureus producing hydroxyl radicals in order to kill the bacterial cells in vitro. In a mouse peritonitis model, FOM treatment increased the bactericidal extra- and intracellular activity in vivo, and FOM strengthened ROS and ET production from peritoneal lavage fluid ex vivo. An IVIS imaging system assay further verified the observed in vivo bactericidal effect of the FOM treatment. This work may provide a deeper understanding of the role of the host’s immune defences and antibiotic interactions in microbial infections. PMID:26778774

Effect of complement Factor H on anti-FHbp serum bactericidal antibody responses of infant rhesus macaques boosted with a licensed meningococcal serogroup B vaccine.

PubMed

Giuntini, Serena; Beernink, Peter T; Granoff, Dan M

2015-12-16

FHbp is a major serogroup B meningococcal vaccine antigen. Binding of complement Factor H (FH) to FHbp is specific for human and some non-human primate FH. In previous studies, FH binding to FHbp vaccines impaired protective anti-FHbp antibody responses. In this study we investigated anti-FHbp antibody responses to a third dose of a licensed serogroup B vaccine (MenB-4C) in infant macaques vaccinated in a previous study with MenB-4C. Six macaques with high binding of FH to FHbp (FH(high)), and six with FH(low) baseline phenotypes, were immunized three months after dose 2. After dose 2, macaques with the FH(low) baseline phenotype had serum anti-FHbp antibodies that enhanced FH binding to FHbp (functionally converting them to a FH(high) phenotype). In this group, activation of the classical complement pathway (C4b deposition) by serum anti-FHbp antibody, and anti-FHbp serum bactericidal titers were lower after dose 3 than after dose 2 (p<0.02). In macaques with the FH(high) baseline phenotype, the respective anti-FHbp C4b deposition and bactericidal titers were similar after doses 2 and 3. Two macaques developed serum anti-FH autoantibodies after dose 2, which were not detected after dose 3. In conclusion, in macaques with the FH(low) baseline phenotype whose post-dose 2 serum anti-FHbp antibodies had converted them to FH(high), the anti-FHbp antibody repertoire to dose 3 was skewed to less protective epitopes than after dose 2. Mutant FHbp vaccines that eliminate FH binding may avoid eliciting anti-FHbp antibodies that enhance FH binding, and confer greater protection with less risk of inducing anti-FH autoantibodies than FHbp vaccines that bind FH. Copyright © 2015 Elsevier Ltd. All rights reserved.

Sublytic complement protects prostate cancer cells from tumour necrosis factor-α-induced cell death.

PubMed

Liu, L; Li, W; Li, Z; Kirschfink, M

2012-08-01

Inflammation is a critical component of tumour progression. Although complement and tumour necrosis factor (TNF)-α potentially exert significant anti-tumour effects, both mediators may also promote tumour progression. It has been demonstrated that sublytic complement confers resistance on tumour cells not only against lytic complement, but also other danger molecules such as perforin. In low concentrations, TNF promotes survival of malignant cells rather than exerting cytotoxic activity. In this study, we tested if sublytic complement is able to interfere with TNF-mediated tumour cell killing. Our results demonstrate that either subcytotoxic concentrations of TNF or sublytic complement rescue prostate carcinoma cells (DU145) from TNF-α-mediated cell death. Upon pretreatment with low-dose TNF-α, but not upon pre-exposure to sublytic complement, TNF resistance was associated with the down-regulation of TNF receptor 1 (TNF-R1) expression. Complement-induced protection against TNF-mediated apoptosis accompanied the induction of anti-apoptotic proteins [B cell leukaemia/lymphoma (Bcl)-2 and Bcl-xL] at an early stage followed by inhibition of the TNF-induced decrease in the amount of Bcl-2 and Bcl-xL. Cell protection also accompanied the inhibition of caspase-8 activation, poly (ADP-ribose) polymerase (PARP)-1 cleavage and the activation of nuclear factor (NF)-κB. Our data extend our current view on the induction of tumour cell resistance against cytotoxic mediators supporting the role of the tumour microenvironment in mediating protection against the anti-cancer immune response. © 2012 The Authors. Clinical and Experimental Immunology © 2012 British Society for Immunology.

Complement C5a Receptor 1 Exacerbates the Pathophysiology of N. meningitidis Sepsis and Is a Potential Target for Disease Treatment

PubMed Central

Herrmann, Johannes B.; Muenstermann, Marcel; Strobel, Lea; Schubert-Unkmeir, Alexandra; Woodruff, Trent M.; Klos, Andreas

2018-01-01

ABSTRACT Sepsis caused by Neisseria meningitidis (meningococcus) is a rapidly progressing, life-threatening disease. Because its initial symptoms are rather unspecific, medical attention is often sought too late, i.e., when the systemic inflammatory response is already unleashed. This in turn limits the success of antibiotic treatment. The complement system is generally accepted as the most important innate immune determinant against invasive meningococcal disease since it protects the host through the bactericidal membrane attack complex. However, complement activation concomitantly liberates the C5a peptide, and it remains unclear whether this potent anaphylatoxin contributes to protection and/or drives the rapidly progressing immunopathogenesis associated with meningococcal disease. Here, we dissected the specific contribution of C5a receptor 1 (C5aR1), the canonical receptor for C5a, using a mouse model of meningococcal sepsis. Mice lacking C3 or C5 displayed susceptibility that was enhanced by >1,000-fold or 100-fold, respectively, consistent with the contribution of these components to protection. In clear contrast, C5ar1−/− mice resisted invasive meningococcal infection and cleared N. meningitidis more rapidly than wild-type (WT) animals. This favorable outcome stemmed from an ameliorated inflammatory cytokine response to N. meningitidis in C5ar1−/− mice in both in vivo and ex vivo whole-blood infections. In addition, inhibition of C5aR1 signaling without interference with the complement bactericidal activity reduced the inflammatory response also in human whole blood. Enticingly, pharmacologic C5aR1 blockade enhanced mouse survival and lowered meningococcal burden even when the treatment was administered after sepsis induction. Together, our findings demonstrate that C5aR1 drives the pathophysiology associated with meningococcal sepsis and provides a promising target for adjunctive therapy. PMID:29362231

Complement C5-inhibiting therapy for the thrombotic microangiopathies: accumulating evidence, but not a panacea

PubMed Central

Brocklebank, Vicky

2017-01-01

Abstract Thrombotic microangiopathy (TMA), characterized by organ injury occurring consequent to severe endothelial damage, can manifest in a diverse range of diseases. In complement-mediated atypical haemolytic uraemic syndrome (aHUS) a primary defect in complement, such as a mutation or autoantibody leading to over activation of the alternative pathway, predisposes to the development of disease, usually following exposure to an environmental trigger. The elucidation of the pathogenesis of aHUS resulted in the successful introduction of the complement inhibitor eculizumab into clinical practice. In other TMAs, although complement activation may be seen, its role in the pathogenesis remains to be confirmed by an interventional trial. Although many case reports in TMAs other than complement-mediated aHUS hint at efficacy, publication bias, concurrent therapies and in some cases the self-limiting nature of disease make broader interpretation difficult. In this article, we will review the evidence for the role of complement inhibition in complement-mediated aHUS and other TMAs. PMID:28980670

Correlation between serum bactericidal activity against Neisseria meningitidis serogroups A, C, W-135 and Y measured using human versus rabbit serum as the complement source.

PubMed Central

CJ, Gill; S, Ram; JA, Welsch; L, DeTora; A, Anemona

2014-01-01

The surrogate of protection against invasive meningococcal disease is the presence of serum bactericidal activity (SBA) at a titer ≥4 in an assay using human serum as the complement source (hSBA). However, for various practical and logistical reasons, many meningococcal vaccines in use today were licensed based on a modified SBA assay that used baby rabbit serum as the complement source (rSBA). To assess the strength of correlation between the two assay systems for serogroups A, C, W-135 and Y, we analyzed a subset of samples from adolescent subjects enrolled in a Phase II study of Novartis’ MenACWY-CRM conjugate vaccine vs. an ACWY polysaccharide vaccine; samples were analyzed in parallel using hSBA and rSBA. We compared geometric mean titers (GMTs), calculated Pearson correlation coefficients between paired hSBA and rSBA results, and calculated sensitivity/specificity and likelihood ratios for an rSBA ≥8 or ≥128 for classifying hSBA ≥4, taking hSBA as the ‘gold standard’. Correlations between hSBA and rSBA ranged from 0.46 to 0.78 for serogroup C, but were weaker for serogroups A, W-135 and Y (range -0.15 to 0.57). In post vaccination samples, nearly all subjects had rSBA titers ≥8, though up to 15% remained seronegative by hSBA. In post vaccination settings, rSBA titers at ≥8 or ≥128 was highly sensitive for an hSBA titer ≥4, but non-specific. In conclusion, results generated by rSBA did not accurately classify serostatus according to hSBA for serogroups A, W-135 and Y. PMID:22075087

Inactivation of a subpopulation of human neutrophils by exposure to ultrahigh-molecular-weight polyethylene wear debris.

PubMed

Bernard, Louis; Vaudaux, Pierre; Huggler, Elzbieta; Stern, Richard; Fréhel, Claude; Francois, Patrice; Lew, Daniel; Hoffmeyer, Pierre

2007-04-01

Polymorphonuclear neutrophils, a first line of defence against invading microbial pathogens, may be attracted by inflammatory mediators triggered by ultrahigh-molecular-weight polyethylene (UHMWPE) wear particles released from orthopaedic prostheses. Phagocytosis of UHMWPE particles by neutrophils may indirectly compromise their phagocytic-bactericidal mechanisms, thus enhancing host susceptibility to microbial infections. In an in vitro assay, pre-exposure of purified human neutrophils to UHMWPE micrometre- and submicrometre-sized wear particles interfered with subsequent Staphylococcos aureus uptake in a heterogeneous way, as assessed by a dual label fluorescence microscopic assay that discriminated intracellular rhodamine-labelled UHMWPE particles from fluorescein isothiocyanate-labelled S. aureus. Indeed, a higher percentage (44%) of neutrophils having engulfed UHMWPE particles lost the ability to phagocytize S. aureus, compared with UHMWPE-free neutrophils (<3%). Pre-exposure of neutrophils to UHMWPE wear particles did not impair but rather stimulated their oxidative burst response in a chemoluminescence assay. The presence of UHMWPE wear particles did not lead to significant overall consumption of complement-mediated opsonic factors nor decreased surface membrane display of neutrophil complement receptors. In conclusion, engulfment of UHMWPE wear particles led to inactivation of S. aureus uptake in nearly half of the neutrophil population, which may potentially impair host clearance mechanisms against pyogenic infections.

Vaccination against Neisseria meningitidis using three variants of the lipoprotein GNA1870.

PubMed

Masignani, Vega; Comanducci, Maurizio; Giuliani, Marzia Monica; Bambini, Stefania; Adu-Bobie, Jeannette; Arico, Beatrice; Brunelli, Brunella; Pieri, Alessandro; Santini, Laura; Savino, Silvana; Serruto, Davide; Litt, David; Kroll, Simon; Welsch, Jo Anne; Granoff, Dan M; Rappuoli, Rino; Pizza, Mariagrazia

2003-03-17

Sepsis and meningitis caused by serogroup B meningococcus are devastating diseases of infants and young adults, which cannot yet be prevented by vaccination. By genome mining, we discovered GNA1870, a new surface-exposed lipoprotein of Neisseria meningitidis that induces high levels of bactericidal antibodies. The antigen is expressed by all strains of N. meningitidis tested. Sequencing of the gene in 71 strains representative of the genetic and geographic diversity of the N. meningitidis population, showed that the protein can be divided into three variants. Conservation within each variant ranges between 91.6 to 100%, while between the variants the conservation can be as low as 62.8%. The level of expression varies between strains, which can be classified as high, intermediate, and low expressors. Antibodies against a recombinant form of the protein elicit complement-mediated killing of the strains that carry the same variant and induce passive protection in the infant rat model. Bactericidal titers are highest against those strains expressing high yields of the protein; however, even the very low expressors are efficiently killed. The novel antigen is a top candidate for the development of a new vaccine against meningococcus.

Screening of Natural Product Derivatives Identifies Two Structurally Related Flavonoids as Potent Quorum Sensing Inhibitors against Gram-Negative Bacteria.

PubMed

Manner, Suvi; Fallarero, Adyary

2018-05-03

Owing to the failure of conventional antibiotics in biofilm control, alternative approaches are urgently needed. Inhibition of quorum sensing (QS) represents an attractive target since it is involved in several processes essential for biofilm formation. In this study, a compound library of natural product derivatives ( n = 3040) was screened for anti-quorum sensing activity using Chromobacterium violaceum as reporter bacteria. Screening assays, based on QS-mediated violacein production and viability, were performed in parallel to identify non-bactericidal QS inhibitors (QSIs). Nine highly active QSIs were identified, while 328 compounds were classified as moderately actives and 2062 compounds as inactives. Re-testing of the highly actives at a lower concentration against C. violaceum , complemented by a literature search, led to the identification of two flavonoid derivatives as the most potent QSIs, and their impact on biofilm maturation in Escherichia coli and Pseudomonas aeruginosa was further investigated. Finally, effects of these leads on swimming and swarming motility of P. aeruginosa were quantified. The identified flavonoids affected all the studied QS-related functions at micromolar concentrations. These compounds can serve as starting points for further optimization and development of more potent QSIs as adjunctive agents used with antibiotics in the treatment of biofilms.

Mesophilic Aeromonas sp. serogroup O:11 resistance to complement-mediated killing.

PubMed Central

Merino, S; Rubires, X; Aguilar, A; Albertí, S; Hernandez-Allés, S; Benedí, V J; Tomas, J M

1996-01-01

The complement activation by and resistance to complement-mediated killing of Aeromonas sp. strains from serogroup O:11 were investigated by using different wild-type strains (with an S-layer characteristic of this serogroup) and their isogenic mutants characterized for their surface components (S-layer and lipopolysaccharide [LPS]). All of the Aeromonas sp. serogroup O:11 wild-type strains are unable to activate complement, which suggested that the S-layer completely covered the LPS molecules. We found that the classical complement pathway is involved in serum killing of susceptible Aeromonas sp. mutant strains of serogroup O11, while the alternative complement pathway seems not to be involved, and that the complement activation seems to be independent of antibody. The smooth mutant strains devoid of the S-layer (S-layer isogenic mutants) or isogenic LPS mutant strains with a complete or rather complete LPS core (also without the S-layer) are able to activate complement but are resistant to complement-mediated killing. The reasons for this resistance are that C3b is rapidly degraded, and therefore the lytic membrane attack complex (C5b-9) is not formed. Isogenic LPS rough mutants with an incomplete LPS core are serum sensitive because they bind more C3b than the resistant strains, the C3b is not completely degraded, and therefore the lytic complex (C5b-9) is formed. PMID:8945581

Eliminating Factor H-Binding Activity of Borrelia burgdorferi CspZ Combined with Virus-Like Particle Conjugation Enhances Its Efficacy as a Lyme Disease Vaccine.

PubMed

Marcinkiewicz, Ashley L; Lieknina, Ilva; Kotelovica, Svetlana; Yang, Xiuli; Kraiczy, Peter; Pal, Utpal; Lin, Yi-Pin; Tars, Kaspars

2018-01-01

The spirochete Borrelia burgdorferi is the causative agent of Lyme disease, the most common tick-borne disease in the US and Europe. No potent human vaccine is currently available. The innate immune complement system is vital to host defense against pathogens, as complement activation on the surface of spirochetes results in bacterial killing. Complement system is inhibited by the complement regulator factor H (FH). To escape killing, B. burgdorferi produces an outer surface protein CspZ that binds FH to inhibit complement activation on the cell surface. Immunization with CspZ alone does not protect mice from infection, which we speculate is because FH-binding cloaks potentially protective epitopes. We modified CspZ by conjugating to virus-like particles (VLP-CspZ) and eliminating FH binding (modified VLP-CspZ) to increase immunogenicity. We observed greater bactericidal antibody titers in mice vaccinated with modified VLP-CspZ: A serum dilution of 1:395 (modified VLP-CspZ) vs 1:143 (VLP-CspZ) yielded 50% borreliacidal activity. Immunizing mice with modified VLP-CspZ cleared spirochete infection, as did passive transfer of elicited antibodies. This work developed a novel Lyme disease vaccine candidate by conjugating CspZ to VLP and eliminating FH-binding ability. Such a strategy of conjugating an antigen to a VLP and eliminating binding to the target ligand can serve as a general model for developing vaccines against other bacterial infectious agents.

Extended low-resolution structure of a Leptospira antigen offers high bactericidal antibody accessibility amenable to vaccine design

PubMed Central

Tseng, Andrew; Suguiura, Igor Massahiro de Souza; McDonough, Sean P; Sritrakul, Tepyuda; Li, Ting; Lin, Yi-Pin; Gillilan, Richard E

2017-01-01

Pathogens rely on proteins embedded on their surface to perform tasks essential for host infection. These obligatory structures exposed to the host immune system provide important targets for rational vaccine design. Here, we use a systematically designed series of multi-domain constructs in combination with small angle X-ray scattering (SAXS) to determine the structure of the main immunoreactive region from a major antigen from Leptospira interrogans, LigB. An anti-LigB monoclonal antibody library exhibits cell binding and bactericidal activity with extensive domain coverage complementing the elongated architecture observed in the SAXS structure. Combining antigenic motifs in a single-domain chimeric immunoglobulin-like fold generated a vaccine that greatly enhances leptospiral protection over vaccination with single parent domains. Our study demonstrates how understanding an antigen’s structure and antibody accessible surfaces can guide the design and engineering of improved recombinant antigen-based vaccines. PMID:29210669

Distinct polymer architecture mediates switching of complement activation pathways at the nanosphere-serum interface: implications for stealth nanoparticle engineering.

PubMed

Hamad, Islam; Al-Hanbali, Othman; Hunter, A Christy; Rutt, Kenneth J; Andresen, Thomas L; Moghimi, S Moein

2010-11-23

Nanoparticles with surface projected polyethyleneoxide (PEO) chains in "mushroom-brush" and "brush" configurations display stealth properties in systemic circulation and have numerous applications in site-specific targeting for controlled drug delivery and release as well as diagnostic imaging. We report on the "structure-activity" relationship pertaining to surface-immobilized PEO of various configurations on model nanoparticles, and the initiation of complement cascade, which is the most ancient component of innate human immunity, and its activation may induce clinically significant adverse reactions in some individuals. Conformational states of surface-projected PEO chains, arising from the block copolymer poloxamine 908 adsorption, on polystyrene nanoparticles trigger complement activation differently. Alteration of copolymer architecture on nanospheres from mushroom to brush configuration not only switches complement activation from C1q-dependent classical to lectin pathway but also reduces the level of generated complement activation products C4d, Bb, C5a, and SC5b-9. Also, changes in adsorbed polymer configuration trigger alternative pathway activation differently and through different initiators. Notably, the role for properdin-mediated activation of alternative pathway was only restricted to particles displaying PEO chains in a transition mushroom-brush configuration. Since nanoparticle-mediated complement activation is of clinical concern, our findings provide a rational basis for improved surface engineering and design of immunologically safer stealth and targetable nanosystems with polymers for use in clinical medicine.

Human Tamm-Horsfall protein, a renal specific protein, serves as a cofactor in complement 3b degradation

PubMed Central

2017-01-01

Tamm-Horsfall protein (THP) is an abundant urinary protein of renal origin. We hypothesize that THP can act as an inhibitor of complement since THP binds complement 1q (C1q) of the classical complement pathway, inhibits activation of this pathway, and is important in decreasing renal ischemia-reperfusion injury (a complement-mediated condition). In this study, we began to investigate whether THP interacted with the alternate complement pathway via complement factor H (CFH). THP was shown to bind CFH using ligand blots and in an ELISA (KD of 1 × 10−6 M). Next, the ability of THP to alter CFH’s normal action as it functioned as a cofactor in complement factor I (CFI)–mediated complement 3b (C3b) degradation was investigated. Unexpectedly, control experiments in these in vitro assays suggested that THP, without added CFH, could act as a cofactor in CFI-mediated C3b degradation. This cofactor activity was present equally in THP isolated from 10 different individuals. While an ELISA demonstrated small amounts of CFH contaminating THP samples, these CFH amounts were insufficient to explain the degree of cofactor activity present in THP. An ELISA demonstrated that THP directly bound C3b (KD ~ 5 × 10−8 m), a prerequisite for a protein acting as a C3b degradation cofactor. The cofactor activity of THP likely resides in the protein portion of THP since partially deglycosylated THP still retained cofactor activity. In conclusion, THP appears to participate directly in complement inactivation by its ability to act as a cofactor for C3b degradation, thus adding support to the hypothesis that THP might act as an endogenous urinary tract inhibitor of complement. PMID:28742158

Subinhibitory quinupristin/dalfopristin attenuates virulence of Staphylococcus aureus.

PubMed

Koszczol, Carmen; Bernardo, Katussevani; Krönke, Martin; Krut, Oleg

2006-09-01

The semi-synthetic streptogramin quinupristin/dalfopristin antibiotic exerts potent bactericidal activity against Staphylococcus aureus. We investigated whether, like other bactericidal antibiotics used at subinhibitory concentrations, quinupristin/dalfopristin enhances release of toxins by Gram-positive cocci. The activity of quinupristin/dalfopristin on exotoxin release by S. aureus was investigated by 2D SDS-PAGE combined with MALDI-TOF/MS analysis and by western blotting. We show that quinupristin/dalfopristin at subinhibitory concentrations reduces the release of S. aureus factors that induce tumour necrosis factor secretion in macrophages. Furthermore, quinupristin/dalfopristin but not linezolid attenuated S. aureus-mediated killing of infected host cells. When added to S. aureus cultures at different stages of bacterial growth, quinupristin/dalfopristin reduced in a dose-dependent manner the release of specific virulence factors (e.g. autolysin, protein A, alpha- and beta-haemolysins, lipases). In contrast, other presumably non-toxic exoproteins remained unchanged. The results of the present study suggest that subinhibitory quinupristin/dalfopristin inhibits virulence factor release by S. aureus, which might be especially helpful for the treatment of S. aureus infections, where both bactericidal as well as anti-toxin activity may be advantageous.

Ulex europaeus agglutinin II (UEA-II) is a novel, potent inhibitor of complement activation.

PubMed

Lekowski, R; Collard, C D; Reenstra, W R; Stahl, G L

2001-02-01

Complement is an important mediator of vascular injury following oxidative stress. We recently demonstrated that complement activation following endothelial oxidative stress is mediated by mannose-binding lectin (MBL) and activation of the lectin complement pathway. Here, we investigated whether nine plant lectins which have a binding profile similar to that of MBL competitively inhibit MBL deposition and subsequent complement activation following human umbilical vein endothelial cell (HUVEC) oxidative stress. HUVEC oxidative stress (1% O(2), 24 hr) significantly increased Ulex europaeus agglutinin II (UEA-II) binding by 72 +/- 9% compared to normoxic cells. UEA-II inhibited MBL binding to HUVEC in a concentration-dependent manner following oxidative stress. Further, MBL inhibited UEA-II binding to HUVEC in a concentration-dependent manner following oxidative stress, suggesting a common ligand. UEA-II (< or = 100 micromol/L) did not attenuate the hemolytic activity, nor did it inhibit C3a des Arg formation from alternative or classical complement pathway-specific hemolytic assays. C3 deposition (measured by ELISA) following HUVEC oxidative stress was inhibited by UEA-II in a concentration-dependent manner (IC(50) = 10 pmol/L). UEA-II inhibited C3 and MBL co-localization (confocal microscopy) in a concentration-dependent manner on HUVEC following oxidative stress (IC(50) approximately 1 pmol/L). Finally, UEA-II significantly inhibited complement-dependent neutrophil chemotaxis, but failed to inhibit fMLP-mediated chemotaxis, following endothelial oxidative stress. These data demonstrate that UEA-II is a novel, potent inhibitor of human MBL deposition and complement activation following human endothelial oxidative stress.

Ulex europaeus agglutinin II (UEA-II) is a novel, potent inhibitor of complement activation

PubMed Central

Lekowski, Robert; Collard, Charles D.; Reenstra, Wende R.; Stahl, Gregory L.

2001-01-01

Complement is an important mediator of vascular injury following oxidative stress. We recently demonstrated that complement activation following endothelial oxidative stress is mediated by mannose-binding lectin (MBL) and activation of the lectin complement pathway. Here, we investigated whether nine plant lectins which have a binding profile similar to that of MBL competitively inhibit MBL deposition and subsequent complement activation following human umbilical vein endothelial cell (HUVEC) oxidative stress. HUVEC oxidative stress (1% O2, 24 hr) significantly increased Ulex europaeus agglutinin II (UEA-II) binding by 72 ± 9% compared to normoxic cells. UEA-II inhibited MBL binding to HUVEC in a concentration-dependent manner following oxidative stress. Further, MBL inhibited UEA-II binding to HUVEC in a concentration-dependent manner following oxidative stress, suggesting a common ligand. UEA-II (≤ 100 μmol/L) did not attenuate the hemolytic activity, nor did it inhibit C3a des Arg formation from alternative or classical complement pathway-specific hemolytic assays. C3 deposition (measured by ELISA) following HUVEC oxidative stress was inhibited by UEA-II in a concentration-dependent manner (IC50 = 10 pmol/L). UEA-II inhibited C3 and MBL co-localization (confocal microscopy) in a concentration-dependent manner on HUVEC following oxidative stress (IC50 ≈ 1 pmol/L). Finally, UEA-II significantly inhibited complement-dependent neutrophil chemotaxis, but failed to inhibit fMLP-mediated chemotaxis, following endothelial oxidative stress. These data demonstrate that UEA-II is a novel, potent inhibitor of human MBL deposition and complement activation following human endothelial oxidative stress. PMID:11266613

Essential Oils as Components of a Diet-Based Approach to Management of Helicobacter Infection

PubMed Central

Bergonzelli, G. E.; Donnicola, D.; Porta, N.; Corthésy-Theulaz, I. E.

2003-01-01

An increased density of Helicobacter pylori in the gastric mucosa can be associated with more severe gastritis and an increased incidence of peptic ulcers. Therefore, people with asymptomatic gastritis would certainly benefit from a nutritional approach to help them manage the infection and therefore decrease the risk of development of associated pathologies. We analyzed the activities of 60 essential oils against H. pylori P1 and identified 30 oils that affected growth, with in vitro inhibition zones ranging between 0.7 and 6.3 cm in diameter. We further analyzed the effects of 16 oils with different activities on H. pylori P1 viability. Fifteen showed strong bactericidal activities, with minimal bactericidal concentrations after 24 h ranging from 0.02 to 0.1 g/liter at pH 7.4. Even though slight variations in activities were observed, the essential oils that displayed the strongest bactericidal potentials against H. pylori P1 were also active against other Helicobacter strains tested. Among the pure constituents of different essential oils tested, carvacrol, isoeugenol, nerol, citral, and sabinene exhibited the strongest anti-H. pylori activities. Although oral treatment of H. pylori SS1-infected mice with carrot seed oil did not result in significant decreases in the bacterial loads in the treated animals compared to those in the control animals, in all experiments performed, the infection was cleared in 20 to 30% of carrot seed oil-treated animals. Our results indicate that essential oils are unlikely to be efficient anti-Helicobacter agents in vivo. However, their effects may not be irrelevant if one plans to use them as food additives to complement present therapies. PMID:14506036

Relative Contribution of Cellular Complement Inhibitors CD59, CD46, and CD55 to Parainfluenza Virus 5 Inhibition of Complement-Mediated Neutralization

PubMed Central

Li, Yujia; Parks, Griffith D.

2018-01-01

The complement system is a part of the innate immune system that viruses need to face during infections. Many viruses incorporate cellular regulators of complement activation (RCA) to block complement pathways and our prior work has shown that Parainfluenza virus 5 (PIV5) incorporates CD55 and CD46 to delay complement-mediated neutralization. In this paper, we tested the role of a third individual RCA inhibitor CD59 in PIV5 interactions with complement pathways. Using a cell line engineered to express CD59, we show that small levels of functional CD59 are associated with progeny PIV5, which is capable of blocking assembly of the C5b-C9 membrane attack complex (MAC). PIV5 containing CD59 (PIV5-CD59) showed increased resistance to complement-mediated neutralization in vitro comparing to PIV5 lacking regulators. Infection of A549 cells with PIV5 and RSV upregulated CD59 expression. TGF-beta treatment of PIV5-infected cells also increased cell surface CD59 expression and progeny virions were more resistant to complement-mediated neutralization. A comparison of individual viruses containing only CD55, CD46, or CD59 showed a potency of inhibiting complement-mediated neutralization, which followed a pattern of CD55 > CD46 > CD59. PMID:29693588

Complement-Mediated Enhancement of Monocyte Adhesion to Endothelial Cells by HLA Antibodies, and Blockade by a Specific Inhibitor of the Classical Complement Cascade, TNT003

PubMed Central

Valenzuela, Nicole M.; Thomas, Kimberly A.; Mulder, Arend; Parry, Graham C.; Panicker, Sandip; Reed, Elaine F.

2017-01-01

Background Antibody-mediated rejection (AMR) of most solid organs is characterized by evidence of complement activation and/or intragraft macrophages (C4d + and CD68+ biopsies). We previously demonstrated that crosslinking of HLA I by antibodies triggered endothelial activation and monocyte adhesion. We hypothesized that activation of the classical complement pathway at the endothelial cell surface by HLA antibodies would enhance monocyte adhesion through soluble split product generation, in parallel with direct endothelial activation downstream of HLA signaling. Methods Primary human aortic endothelial cells (HAEC) were stimulated with HLA class I antibodies in the presence of intact human serum complement. C3a and C5a generation, endothelial P-selectin expression, and adhesion of human primary and immortalized monocytes (Mono Mac 6) were measured. Alternatively, HAEC or monocytes were directly stimulated with purified C3a or C5a. Classical complement activation was inhibited by pretreatment of complement with an anti-C1s antibody (TNT003). Results Treatment of HAEC with HLA antibody and human complement increased the formation of C3a and C5a. Monocyte recruitment by human HLA antibodies was enhanced in the presence of intact human serum complement or purified C3a or C5a. Specific inhibition of the classical complement pathway using TNT003 or C1q-depleted serum significantly reduced adhesion of monocytes in the presence of human complement. Conclusions Despite persistent endothelial viability in the presence of HLA antibodies and complement, upstream complement anaphylatoxin production exacerbates endothelial exocytosis and leukocyte recruitment. Upstream inhibition of classical complement may be therapeutic to dampen mononuclear cell recruitment and endothelial activation characteristic of microvascular inflammation during AMR. PMID:28640789

Shape-dependent bactericidal activity of copper oxide nanoparticle mediated by DNA and membrane damage

DOE Office of Scientific and Technical Information (OSTI.GOV)

Laha, Dipranjan; Pramanik, Arindam; Laskar, Aparna

Highlights: • Spherical and sheet shaped copper oxide nanoparticles were synthesized. • Physical characterizations of these nanoparticles were done by TEM, DLS, XRD, FTIR. • They showed shape dependent antibacterial activity on different bacterial strain. • They induced both membrane damage and ROS mediated DNA damage in bacteria. - Abstract: In this work, we synthesized spherical and sheet shaped copper oxide nanoparticles and their physical characterizations were done by the X-ray diffraction, fourier transform infrared spectroscopy, transmission electron microscopy and dynamic light scattering. The antibacterial activity of these nanoparticles was determined on both gram positive and gram negative bacterial. Sphericalmore » shaped copper oxide nanoparticles showed more antibacterial property on gram positive bacteria where as sheet shaped copper oxide nanoparticles are more active on gram negative bacteria. We also demonstrated that copper oxide nanoparticles produced reactive oxygen species in both gram negative and gram positive bacteria. Furthermore, they induced membrane damage as determined by atomic force microscopy and scanning electron microscopy. Thus production of and membrane damage are major mechanisms of the bactericidal activity of these copper oxide nanoparticles. Finally it was concluded that antibacterial activity of nanoparticles depend on physicochemical properties of copper oxide nanoparticles and bacterial strain.« less

IgG Fc domains that bind C1q but not effector Fcγ receptors delineate the importance of complement-mediated effector functions.

PubMed

Lee, Chang-Han; Romain, Gabrielle; Yan, Wupeng; Watanabe, Makiko; Charab, Wissam; Todorova, Biliana; Lee, Jiwon; Triplett, Kendra; Donkor, Moses; Lungu, Oana I; Lux, Anja; Marshall, Nicholas; Lindorfer, Margaret A; Goff, Odile Richard-Le; Balbino, Bianca; Kang, Tae Hyun; Tanno, Hidetaka; Delidakis, George; Alford, Corrine; Taylor, Ronald P; Nimmerjahn, Falk; Varadarajan, Navin; Bruhns, Pierre; Zhang, Yan Jessie; Georgiou, George

2017-08-01

Engineered crystallizable fragment (Fc) regions of antibody domains, which assume a unique and unprecedented asymmetric structure within the homodimeric Fc polypeptide, enable completely selective binding to the complement component C1q and activation of complement via the classical pathway without any concomitant engagement of the Fcγ receptor (FcγR). We used the engineered Fc domains to demonstrate in vitro and in mouse models that for therapeutic antibodies, complement-dependent cell-mediated cytotoxicity (CDCC) and complement-dependent cell-mediated phagocytosis (CDCP) by immunological effector molecules mediated the clearance of target cells with kinetics and efficacy comparable to those of the FcγR-dependent effector functions that are much better studied, while they circumvented certain adverse reactions associated with FcγR engagement. Collectively, our data highlight the importance of CDCC and CDCP in monoclonal-antibody function and provide an experimental approach for delineating the effect of complement-dependent effector-cell engagement in various therapeutic settings.

Human Salivary Protein Histatin 5 Has Potent Bactericidal Activity against ESKAPE Pathogens

PubMed Central

Du, Han; Puri, Sumant; McCall, Andrew; Norris, Hannah L.; Russo, Thomas; Edgerton, Mira

2017-01-01

ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumanni, Pseudomonas aeruginosa, and Enterobacter species) pathogens have characteristic multiple-drug resistance and cause an increasing number of nosocomial infections worldwide. Peptide-based therapeutics to treat ESKAPE infections might be an alternative to conventional antibiotics. Histatin 5 (Hst 5) is a salivary cationic histidine-rich peptide produced only in humans and higher primates. It has high antifungal activity against Candida albicans through an energy-dependent, non-lytic process; but its bactericidal effects are less known. We found Hst 5 has bactericidal activity against S. aureus (60–70% killing) and A. baumannii (85–90% killing) in 10 and 100 mM sodium phosphate buffer (NaPB), while killing of >99% of P. aeruginosa, 60–80% E. cloacae and 20–60% of E. faecium was found in 10 mM NaPB. Hst 5 killed 60% of biofilm cells of P. aeruginosa, but had reduced activity against biofilms of S. aureus and A. baumannii. Hst 5 killed 20% of K. pneumonia biofilm cells but not planktonic cells. Binding and uptake studies using FITC-labeled Hst 5 showed E. faecium and E. cloacae killing required Hst 5 internalization and was energy dependent, while bactericidal activity was rapid against P. aeruginosa and A. baumannii suggesting membrane disruption. Hst 5-mediated killing of S. aureus was both non-lytic and energy independent. Additionally, we found that spermidine conjugated Hst 5 (Hst5-Spd) had improved killing activity against E. faecium, E. cloacae, and A. baumannii. Hst 5 or its derivative has antibacterial activity against five out of six ESKAPE pathogens and may be an alternative treatment for these infections. PMID:28261570

Human Salivary Protein Histatin 5 Has Potent Bactericidal Activity against ESKAPE Pathogens.

PubMed

Du, Han; Puri, Sumant; McCall, Andrew; Norris, Hannah L; Russo, Thomas; Edgerton, Mira

2017-01-01

ESKAPE ( Enterococcus faecium , Staphylococcus aureus , Klebsiella pneumoniae , Acinetobacter baumanni , Pseudomonas aeruginosa , and Enterobacter species) pathogens have characteristic multiple-drug resistance and cause an increasing number of nosocomial infections worldwide. Peptide-based therapeutics to treat ESKAPE infections might be an alternative to conventional antibiotics. Histatin 5 (Hst 5) is a salivary cationic histidine-rich peptide produced only in humans and higher primates. It has high antifungal activity against Candida albicans through an energy-dependent, non-lytic process; but its bactericidal effects are less known. We found Hst 5 has bactericidal activity against S. aureus (60-70% killing) and A. baumannii (85-90% killing) in 10 and 100 mM sodium phosphate buffer (NaPB), while killing of >99% of P. aeruginosa , 60-80% E. cloacae and 20-60% of E. faecium was found in 10 mM NaPB. Hst 5 killed 60% of biofilm cells of P. aeruginosa , but had reduced activity against biofilms of S. aureus and A. baumannii . Hst 5 killed 20% of K. pneumonia biofilm cells but not planktonic cells. Binding and uptake studies using FITC-labeled Hst 5 showed E. faecium and E. cloacae killing required Hst 5 internalization and was energy dependent, while bactericidal activity was rapid against P. aeruginosa and A. baumannii suggesting membrane disruption. Hst 5-mediated killing of S. aureus was both non-lytic and energy independent. Additionally, we found that spermidine conjugated Hst 5 (Hst5-Spd) had improved killing activity against E. faecium, E. cloacae , and A. baumannii . Hst 5 or its derivative has antibacterial activity against five out of six ESKAPE pathogens and may be an alternative treatment for these infections.

Complement activation by carbon nanotubes and its influence on the phagocytosis and cytokine response by macrophages.

PubMed

Pondman, Kirsten M; Sobik, Martin; Nayak, Annapurna; Tsolaki, Anthony G; Jäkel, Anne; Flahaut, Emmanuel; Hampel, Silke; Ten Haken, Bennie; Sim, Robert B; Kishore, Uday

2014-08-01

Carbon nanotubes (CNTs) have promised a range of applications in biomedicine. Although influenced by the dispersants used, CNTs are recognized by the innate immune system, predominantly by the classical pathway of the complement system. Here, we confirm that complement activation by the CNT used continues up to C3 and C5, indicating that the entire complement system is activated including the formation of membrane-attack complexes. Using recombinant forms of the globular regions of human C1q (gC1q) as inhibitors of CNT-mediated classical pathway activation, we show that C1q, the first recognition subcomponent of the classical pathway, binds CNTs via the gC1q domain. Complement opsonisation of CNTs significantly enhances their uptake by U937 cells, with concomitant downregulation of pro-inflammatory cytokines and up-regulation of anti-inflammatory cytokines in both U937 cells and human monocytes. We propose that CNT-mediated complement activation may cause recruitment of cellular infiltration, followed by phagocytosis without inducing a pro-inflammatory immune response. This study highlights the importance of the complement system in response to carbon nanontube administration, suggesting that the ensuing complement activation may cause recruitment of cellular infiltration, followed by phagocytosis without inducing a pro-inflammatory immune response. Copyright © 2014 Elsevier Inc. All rights reserved.

Enhancement of neutrophil autophagy by an IVIG preparation against multidrug-resistant bacteria as well as drug-sensitive strains.

PubMed

Itoh, Hiroshi; Matsuo, Hidemasa; Kitamura, Naoko; Yamamoto, Sho; Higuchi, Takeshi; Takematsu, Hiromu; Kamikubo, Yasuhiko; Kondo, Tadakazu; Yamashita, Kouhei; Sasada, Masataka; Takaori-Kondo, Akifumi; Adachi, Souichi

2015-07-01

Autophagy occurs in human neutrophils after the phagocytosis of multidrug-resistant bacteria and drug-sensitive strains, including Escherichia coli and Pseudomonas aeruginosa. The present study detected autophagy by immunoblot analysis of LC3B conversion, by confocal scanning microscopic examination of LC3B aggregate formation and by transmission electron microscopic examination of bacteria-containing autophagosomes. Patients with severe bacterial infections are often treated with IVIG alongside antimicrobial agents. Here, we showed that IVIG induced neutrophil-mediated phagocytosis of multidrug-resistant strains. Compared with untreated neutrophils, neutrophils exposed to IVIG showed increased levels of bacterial cell killing, phagocytosis, O(2)(-) release, MPO release, and NET formation. IVIG also increased autophagy in these cells. Inhibiting the late phase of autophagy (fusion of lysosomes with autophagosomes) with bafilomycin A1-reduced, neutrophil-mediated bactericidal activity. These findings indicate that autophagy plays a critical role in the bactericidal activity mediated by human neutrophils. Furthermore, the autophagosomes within the neutrophils contained bacteria only and their organelles only, or both bacteria and their organelles, a previously undocumented observation. Taken together, these results suggest that the contents of neutrophil autophagosomes may be derived from specific autophagic systems, which provide the neutrophil with an advantage. Thus, IVIG promotes the neutrophil-mediated killing of multidrug-resistant bacteria as well as drug-sensitive strains. © Society for Leukocyte Biology.

Enhancement of neutrophil autophagy by an IVIG preparation against multidrug-resistant bacteria as well as drug-sensitive strains

PubMed Central

Itoh, Hiroshi; Matsuo, Hidemasa; Kitamura, Naoko; Yamamoto, Sho; Higuchi, Takeshi; Takematsu, Hiromu; Kamikubo, Yasuhiko; Kondo, Tadakazu; Yamashita, Kouhei; Sasada, Masataka; Takaori-Kondo, Akifumi; Adachi, Souichi

2015-01-01

Autophagy occurs in human neutrophils after the phagocytosis of multidrug-resistant bacteria and drug-sensitive strains, including Escherichia coli and Pseudomonas aeruginosa. The present study detected autophagy by immunoblot analysis of LC3B conversion, by confocal scanning microscopic examination of LC3B aggregate formation and by transmission electron microscopic examination of bacteria-containing autophagosomes. Patients with severe bacterial infections are often treated with IVIG alongside antimicrobial agents. Here, we showed that IVIG induced neutrophil-mediated phagocytosis of multidrug-resistant strains. Compared with untreated neutrophils, neutrophils exposed to IVIG showed increased levels of bacterial cell killing, phagocytosis, O2− release, MPO release, and NET formation. IVIG also increased autophagy in these cells. Inhibiting the late phase of autophagy (fusion of lysosomes with autophagosomes) with bafilomycin A1-reduced, neutrophil-mediated bactericidal activity. These findings indicate that autophagy plays a critical role in the bactericidal activity mediated by human neutrophils. Furthermore, the autophagosomes within the neutrophils contained bacteria only and their organelles only, or both bacteria and their organelles, a previously undocumented observation. Taken together, these results suggest that the contents of neutrophil autophagosomes may be derived from specific autophagic systems, which provide the neutrophil with an advantage. Thus, IVIG promotes the neutrophil-mediated killing of multidrug-resistant bacteria as well as drug-sensitive strains. PMID:25908735

An Anti-C1s Monoclonal, TNT003, Inhibits Complement Activation Induced by Antibodies Against HLA.

PubMed

Thomas, K A; Valenzuela, N M; Gjertson, D; Mulder, A; Fishbein, M C; Parry, G C; Panicker, S; Reed, E F

2015-08-01

Antibody-mediated rejection (AMR) of solid organ transplants (SOT) is characterized by damage triggered by donor-specific antibodies (DSA) binding donor Class I and II HLA (HLA-I and HLA-II) expressed on endothelial cells. While F(ab')2 portions of DSA cause cellular activation and proliferation, Fc regions activate the classical complement cascade, resulting in complement deposition and leukocyte recruitment, both hallmark features of AMR. We characterized the ability of an anti-C1s monoclonal antibody, TNT003, to inhibit HLA antibody (HLA-Ab)-induced complement activation. Complement deposition induced by HLA-Ab was evaluated using novel cell- and bead-based assays. Human aortic endothelial cells (HAEC) were cultured with HLA-Ab and human complement; production of activated complement proteins was measured by flow cytometry. Additionally, C3d deposition was measured on single antigen beads (SAB) mixed with HLA-Ab and human complement. TNT003 inhibited HLA-Ab mediated complement deposition on HAEC in a concentration-dependent manner; C3a, C4a and C5a anaphylatoxin production was also diminished by TNT003. Finally, TNT003 blocked C3d deposition induced by Class I (HLAI-Ab)- and Class II (HLAII-Ab)-specific antibodies on SAB. These data suggest TNT003 may be useful for modulating the effects of DSA, as TNT003 inhibits complement deposition and split product formation generated by HLA-I/II-Ab in vitro. © 2015 The Authors. American Journal of Transplantation Published by Wiley Periodicals, Inc.

An Anti-C1s Monoclonal, TNT003, Inhibits Complement Activation Induced by Antibodies Against HLA

PubMed Central

Thomas, K A; Valenzuela, N M; Gjertson, D; Mulder, A; Fishbein, M C; Parry, G C; Panicker, S; Reed, E F

2015-01-01

Antibody-mediated rejection (AMR) of solid organ transplants (SOT) is characterized by damage triggered by donor-specific antibodies (DSA) binding donor Class I and II HLA (HLA-I and HLA-II) expressed on endothelial cells. While F(ab′)2 portions of DSA cause cellular activation and proliferation, Fc regions activate the classical complement cascade, resulting in complement deposition and leukocyte recruitment, both hallmark features of AMR. We characterized the ability of an anti-C1s monoclonal antibody, TNT003, to inhibit HLA antibody (HLA-Ab)-induced complement activation. Complement deposition induced by HLA-Ab was evaluated using novel cell- and bead-based assays. Human aortic endothelial cells (HAEC) were cultured with HLA-Ab and human complement; production of activated complement proteins was measured by flow cytometry. Additionally, C3d deposition was measured on single antigen beads (SAB) mixed with HLA-Ab and human complement. TNT003 inhibited HLA-Ab mediated complement deposition on HAEC in a concentration-dependent manner; C3a, C4a and C5a anaphylatoxin production was also diminished by TNT003. Finally, TNT003 blocked C3d deposition induced by Class I (HLAI-Ab)- and Class II (HLAII-Ab)-specific antibodies on SAB. These data suggest TNT003 may be useful for modulating the effects of DSA, as TNT003 inhibits complement deposition and split product formation generated by HLA-I/II-Ab in vitro. PMID:25904443

Transient Receptor Potential Channel 6 (TRPC6) Protects Podocytes during Complement-mediated Glomerular Disease*

PubMed Central

Kistler, Andreas D.; Singh, Geetika; Altintas, Mehmet M.; Yu, Hao; Fernandez, Isabel C.; Gu, Changkyu; Wilson, Cory; Srivastava, Sandeep Kumar; Dietrich, Alexander; Walz, Katherina; Kerjaschki, Dontscho; Ruiz, Phillip; Dryer, Stuart; Sever, Sanja; Dinda, Amit K.; Faul, Christian; Reiser, Jochen

2013-01-01

Gain-of-function mutations in the calcium channel TRPC6 lead to autosomal dominant focal segmental glomerulosclerosis and podocyte expression of TRPC6 is increased in some acquired human glomerular diseases, particularly in membranous nephropathy. These observations led to the hypothesis that TRPC6 overactivation is deleterious to podocytes through pathological calcium signaling, both in genetic and acquired diseases. Here, we show that the effects of TRPC6 on podocyte function are context-dependent. Overexpression of TRPC6 alone did not directly affect podocyte morphology and cytoskeletal structure. Unexpectedly, however, overexpression of TRPC6 protected podocytes from complement-mediated injury, whereas genetic or pharmacological TRPC6 inactivation increased podocyte susceptibility to complement. Mechanistically, this effect was mediated by Ca2+/calmodulin-dependent protein kinase II (CaMKII) activation. Podocyte-specific TRPC6 transgenic mice showed stronger CaMKII activation, reduced podocyte foot process effacement and reduced levels of proteinuria during nephrotoxic serum nephritis, whereas TRPC6 null mice exhibited reduced CaMKII activation and higher levels of proteinuria compared with wild type littermates. Human membranous nephropathy biopsy samples showed podocyte staining for active CaMKII, which correlated with the degree of TRPC6 expression. Together, these data suggest a dual and context dependent role of TRPC6 in podocytes where acute activation protects from complement-mediated damage, but chronic overactivation leads to focal segmental glomerulosclerosis. PMID:24194522

Infergen Stimulated Macrophages Restrict Mycobacterium tuberculosis Growth by Autophagy and Release of Nitric Oxide

PubMed Central

Pahari, Susanta; Khan, Nargis; Aqdas, Mohammad; Negi, Shikha; Kaur, Jagdeep; Agrewala, Javed N.

2016-01-01

IFN alfacon-1 (Infergen) is a synthetic form of Interferon (IFN)-α2b. Infergen has immunomodulatory activity and is effective against hepatitis C virus. However, the effect of Infergen (IFG) on Mycobacterium tuberculosis (Mtb) has not yet been reported. Therefore, for the first time, we have studied the influence of IFG in constraining the survival of Mtb in human macrophages. We observed that IFG significantly enhanced the maturation and activation of macrophages. Further, it substantially augmented the secretion of IL-6, nitric oxide (NO) and antigen uptake. Moreover, macrophages exhibited remarkably higher bactericidal activity, as evidenced by reduction in the Mtb growth. Infergen-mediated mechanism was different from the type-1 interferons; since it worked through the activation of NF-κB, phosphorylation of STAT-3 and Akt-PI3K that improved the bactericidal activity through autophagy and NO release. In future, IFG immunotherapy can be a novel strategy for treating patients and controlling TB. PMID:28000752

RecA Inhibitors Potentiate Antibiotic Activity and Block Evolution of Antibiotic Resistance.

PubMed

Alam, Md Kausar; Alhhazmi, Areej; DeCoteau, John F; Luo, Yu; Geyer, C Ronald

2016-03-17

Antibiotic resistance arises from the maintenance of resistance mutations or genes acquired from the acquisition of adaptive de novo mutations or the transfer of resistance genes. Antibiotic resistance is acquired in response to antibiotic therapy by activating SOS-mediated DNA repair and mutagenesis and horizontal gene transfer pathways. Initiation of the SOS pathway promotes activation of RecA, inactivation of LexA repressor, and induction of SOS genes. Here, we have identified and characterized phthalocyanine tetrasulfonic acid RecA inhibitors that block antibiotic-induced activation of the SOS response. These inhibitors potentiate the activity of bactericidal antibiotics, including members of the quinolone, β-lactam, and aminoglycoside families in both Gram-negative and Gram-positive bacteria. They reduce the ability of bacteria to acquire antibiotic resistance mutations and to transfer mobile genetic elements conferring resistance. This study highlights the advantage of including RecA inhibitors in bactericidal antibiotic therapies and provides a new strategy for prolonging antibiotic shelf life. Copyright © 2016 Elsevier Ltd. All rights reserved.

Binding of Free and Immune Complex-Associated Hepatitis C Virus to Erythrocytes Is Mediated by the Complement System.

PubMed

Salam, Kazi Abdus; Wang, Richard Y; Grandinetti, Teresa; De Giorgi, Valeria; Alter, Harvey J; Allison, Robert D

2018-05-09

Erythrocytes bind circulating immune complexes (IC) and facilitate IC clearance from the circulation. Chronic hepatitis C virus (HCV) infection is associated with IC-related disorders. In this study we investigated the kinetics and mechanism of HCV and HCV-IC binding to and dissociation from erythrocytes. Cell culture-produced HCV was mixed with erythrocytes from healthy blood donors and erythrocyte-associated virus particles were quantified. Purified complement proteins, complement-depleted serum, and complement receptor antibodies were used to investigate complement-mediated HCV-erythrocyte binding. Purified HCV-specific immunoglobulin G from a chronic HCV-infected patient was used to study complement-mediated HCV-IC-erythrocyte binding. Binding of HCV to erythrocytes increased 200 to 1,000 fold after adding complement active human serum in the absence of antibody. Opsonization of free HCV occurred within 10 minutes and peak binding to erythrocytes was observed at 20-30 minutes. Complement protein C1 was required for binding, while C2, C3 and C4 significantly enhanced binding. Complement receptor 1 (CR1, CD35) antibodies blocked the binding of HCV to erythrocytes isolated from chronically infected HCV patients and healthy blood donors. HCV-ICs significantly enhanced complement-mediated binding to erythrocytes compared to unbound HCV. Dissociation of complement-opsonized HCV from erythrocytes depended on the presence of Factor I. HCV released by Factor I bound preferentially to CD19+ B cells compared to other leukocytes. These results demonstrate that complement mediates the binding of free and IC-associated HCV to CR1 on erythrocytes, and provide a mechanistic rationale for investigating the differential phenotypic expression of HCV-IC-related disease. This article is protected by copyright. All rights reserved. © 2018 by the American Association for the Study of Liver Diseases.

Factor H: A Complement Regulator in Health and Disease, and a Mediator of Cellular Interactions

PubMed Central

Kopp, Anne; Hebecker, Mario; Svobodová, Eliška; Józsi, Mihály

2012-01-01

Complement is an essential part of innate immunity as it participates in host defense against infections, disposal of cellular debris and apoptotic cells, inflammatory processes and modulation of adaptive immune responses. Several soluble and membrane-bound regulators protect the host from the potentially deleterious effects of uncontrolled and misdirected complement activation. Factor H is a major soluble regulator of the alternative complement pathway, but it can also bind to host cells and tissues, protecting them from complement attack. Interactions of factor H with various endogenous ligands, such as pentraxins, extracellular matrix proteins and DNA are important in limiting local complement-mediated inflammation. Impaired regulatory as well as ligand and cell recognition functions of factor H, caused by mutations or autoantibodies, are associated with the kidney diseases: atypical hemolytic uremic syndrome and dense deposit disease and the eye disorder: age-related macular degeneration. In addition, factor H binds to receptors on host cells and is involved in adhesion, phagocytosis and modulation of cell activation. In this review we discuss current concepts on the physiological and pathophysiological roles of factor H in light of new data and recent developments in our understanding of the versatile roles of factor H as an inhibitor of complement activation and inflammation, as well as a mediator of cellular interactions. A detailed knowledge of the functions of factor H in health and disease is expected to unravel novel therapeutic intervention possibilities and to facilitate the development or improvement of therapies. PMID:24970127

Decreased complement mediated binding of antibody//sup 3/-dsDNA immune complexes to the red blood cells of patients with systemic lupus erythematosus, rheumatoid arthritis, and hematologic malignancies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Taylor, R.P.; Horgan, C.; Buschbacher, R.

1983-06-01

The complement mediated binding of prepared antibody//sup 3/H-dsDNA immune complexes to the red blood cells obtained from a number of patient populations has been investigated. Patients with solid tumors have binding activity similar to that seen in a normal group of individuals. However, a significant fraction of patients with systemic lupus erythematosus, rheumatoid arthritis, and hematologic malignancies have lowered binding activity compared with normal subjects. Quantitative studies indicate the lowered activity probably arises due to a decrease in complement receptors on the respective red blood cells. The potential importance and implications of these findings are briefly discussed.

Susceptibility of pathogenic and nonpathogenic Naegleria ssp

DOE Office of Scientific and Technical Information (OSTI.GOV)

Whiteman, L.Y.

1988-01-01

The susceptibility of four species of Naegleria amoebae to complement-mediated lysis was determined. The amoebicidal activity of normal human serum (NHS) and normal guinea pig serum (NGPS) for Naegleria amoebae was measured by an in vitro cytotoxicity assay. Release of radioactivity from amoebae labeled with {sup 3}H-uridine and visual observation with a compound microscope were used as indices of lysis. Susceptibility or resistance to complement-mediated lysis in vitro correlated with the in vivo pathogenic potential. Nonpathogenic Naegleria amoebae were lysed at a faster rate and at higher cell concentrations than were pathogenic amoebae. Electrophoretic analysis of NHS incubated with pathogenicmore » or nonpathogenic Naegleria spp. demonstrated that amoebae activate the complement cascade resulting in the production of C3 and C5 complement cleavage products. Treatment with papain or trypsin for 1 h, but not with sialidase, increase the susceptibility of highly pathogenic, mouse-passaged N. fowleri to lysis. Treatment with actinomycin D, cycloheximide or various protease inhibitors for 4 h did not increase susceptibility to lysis. Neither a repair process involving de novo protein synthesis nor a complement-inactivating protease appear to account for the increase resistance of N. fowleri amoebae to complement-mediated lysis. A binding study with {sup 125}I radiolabeled C9 indicated that the terminal complement component does not remain stably bound to the membrane of pathogenic amoebae.« less

In Vitro Evaluation of CBR-2092, a Novel Rifamycin-Quinolone Hybrid Antibiotic: Microbiology Profiling Studies with Staphylococci and Streptococci ▿

PubMed Central

Robertson, Gregory T.; Bonventre, Eric J.; Doyle, Timothy B.; Du, Qun; Duncan, Leonard; Morris, Timothy W.; Roche, Eric D.; Yan, Dalai; Lynch, A. Simon

2008-01-01

We present data from antimicrobial assays performed in vitro that pertain to the potential clinical utility of a novel rifamycin-quinolone hybrid antibiotic, CBR-2092, for the treatment of infections mediated by gram-positive cocci. The MIC90s for CBR-2092 against 300 clinical isolates of staphylococci and streptococci ranged from 0.008 to 0.5 μg/ml. Against Staphylococcus aureus, CBR-2092 exhibited prolonged postantibiotic effects (PAEs) and sub-MIC effects (SMEs), with values of 3.2, 6.5, and >8.5 h determined for the PAE (3× MIC), SME (0.12× MIC), and PAE-SME (3× MIC/0.12× MIC) periods, respectively. Studies of genetically defined mutants of S. aureus indicate that CBR-2092 is not a substrate for the NorA or MepA efflux pumps. In minimal bactericidal concentration and time-kill studies, CBR-2092 exhibited bactericidal activity against staphylococci that was retained against rifampin- or intermediate quinolone-resistant strains, with apparent paradoxical cidal characteristics against rifampin-resistant strains. In spontaneous resistance studies, CBR-2092 exhibited activity consistent with balanced contributions from its composite pharmacophores, with a mutant prevention concentration of 0.12 μg/ml and a resistance frequency of <10−12 determined at 1 μg/ml in agar for S. aureus. Similarly, CBR-2092 suppressed the emergence of preexisting rifamycin resistance in time-kill studies undertaken at a high cell density. In studies of the intracellular killing of S. aureus, CBR-2092 exhibited prolonged bactericidal activity that was superior to the activities of moxifloxacin, rifampin, and a cocktail of moxifloxacin and rifampin. Overall, CBR-2092 exhibited promising activity in a range of antimicrobial assays performed in vitro that pertain to properties relevant to the effective treatment of serious infections mediated by gram-positive cocci. PMID:18443106

Two Major Medicinal Honeys Have Different Mechanisms of Bactericidal Activity

PubMed Central

Kwakman, Paulus H. S.; te Velde, Anje A.; de Boer, Leonie; Vandenbroucke-Grauls, Christina M. J. E.; Zaat, Sebastian A. J.

2011-01-01

Honey is increasingly valued for its antibacterial activity, but knowledge regarding the mechanism of action is still incomplete. We assessed the bactericidal activity and mechanism of action of Revamil® source (RS) honey and manuka honey, the sources of two major medical-grade honeys. RS honey killed Bacillus subtilis, Escherichia coli and Pseudomonas aeruginosa within 2 hours, whereas manuka honey had such rapid activity only against B. subtilis. After 24 hours of incubation, both honeys killed all tested bacteria, including methicillin-resistant Staphylococcus aureus, but manuka honey retained activity up to higher dilutions than RS honey. Bee defensin-1 and H2O2 were the major factors involved in rapid bactericidal activity of RS honey. These factors were absent in manuka honey, but this honey contained 44-fold higher concentrations of methylglyoxal than RS honey. Methylglyoxal was a major bactericidal factor in manuka honey, but after neutralization of this compound manuka honey retained bactericidal activity due to several unknown factors. RS and manuka honey have highly distinct compositions of bactericidal factors, resulting in large differences in bactericidal activity. PMID:21394213

Effects of freezing on the bactericidal activity of human milk.

PubMed

Takci, Sahin; Gulmez, Dolunay; Yigit, Sule; Dogan, Ozlem; Dik, Kezban; Hascelik, Gulsen

2012-08-01

Storage of human milk by freezing has been recommended for long-term storage. The present study analyzed the bactericidal activity of human milk on Escherichia coli and Pseudomonas aeruginosa and determined the changes in bactericidal activity following freezing at -20°C and -80°C for 1 month and 3 months. Forty-eight milk samples were collected from 48 lactating mothers. Each sample was divided into 10 aliquots. Two of the samples were processed immediately and the others were stored at both -20°C and -80°C until analysis after 1 month and 3 months of freezing. All of the fresh milk samples showed bactericidal activity against E coli and P aeruginosa. Freezing at -20°C for 1 month did not cause statistically significant alteration in bactericidal activity (P > 0.017), whereas storage for 3 months lowered the degree of bactericidal activity significantly (P < 0.017) against E coli. Bactericidal activity was protected when the samples were stored at -80°C. There was no statistically significant difference in the bactericidal activity of human milk against E coli between freezing at -20°C and -80°C for 1 month (P > 0.017); however, when milk was stored for 3 months, -80°C was significantly more protective (P < 0.017). Freezing at -20°C and -80°C for 1 month and 3 months did not cause any significant change in bactericidal activity against P aeruginosa (P > 0.05). Storage by freezing at -80°C is more appropriate to keep bactericidal capacity of stored human milk >1 month if affordable and available, especially in intensive care settings.

Review article: the potential mechanisms of action of rifaximin in the management of inflammatory bowel diseases.

PubMed

Sartor, R B

2016-01-01

Gut microbiota dysbiosis contributes to the pathogenesis of inflammatory bowel diseases (IBD). Although the microbiota's role in IBD pathogenesis, specifically Crohn's disease (CD), provides a rationale for antibiotic treatment, antibiotic use in CD remains controversial. Rifaximin, traditionally identified as a nonsystemic bactericidal antibiotic, may be therapeutically beneficial for inducing CD remission. To examine the role of rifaximin in the management of IBD and its potential mechanisms of action. A literature search using the following strategy: ('inflammatory bowel disease' OR 'Crohn's' OR 'ulcerative'), 'rifaximin' AND ('barrier' OR 'translocation' OR 'adhesion' OR 'internalization' OR 'pregnane X'), AND 'pregnane X' AND ('Crohn's' OR 'ulcerative colitis' OR 'inflammatory bowel disease'). In vitro data suggest rifaximin mediates changes in epithelial cell physiology and reduces bacterial attachment and internalisation. In experimental colitis models, rifaximin antagonised the effects of tumour necrosis factor-α on intestinal epithelial cells by activating pregnane X receptor, which inhibits nuclear factor-κB-mediated proinflammatory mediators and induces detoxification genes (e.g. multidrug resistance 1 and cytochrome P450 3A4). Rifaximin also inhibits bacterial translocation into the mesenteric lymph nodes. Accumulating evidence suggests that mechanisms of action of rifaximin in IBD may not be limited to direct bactericidal activity; therefore, rifaximin could potentially be redefined as a gut environment modulator. © 2015 John Wiley & Sons Ltd.

Bactericidal activity of wasabi (Wasabia japonica) against Helicobacter pylori.

PubMed

Shin, Il Shik; Masuda, Hideki; Naohide, Kinae

2004-08-01

In this study, the bactericidal activity of Korean and Japanese wasabi roots, stems and leaves against Helicobacter pylori were examined. Allyl isothiocyanate (AIT) in roots, stems and leaves of Korean wasabi were 0.75, 0.18 and 0.32 mg/g, respectively. AIT in roots, stems and leaves of Japanese wasabi were 1.18, 0.41 and 0.38 mg/g, respectively. All parts of wasabi showed bactericidal activities against H. pylori strain NCTC 11637, YS 27 and YS 50. The leaves of both wasabi showed the highest bactericidal activities with the minimum bactericidal concentration of 1.05-1.31 mg of dry weight/ml against three strains of H. pylori. The roots showed a little lower bactericidal activity with 2.09-4.17 mg of dry weight/ml against them. The main component related to antimicrobial activity in wasabi is well known to be AIT. In this study, the bactericidal activity of leaves was higher than that of roots, although AIT amount of leaves was lower than that of roots. These results suggest that certain components besides AIT in wasabi are effective in killing H. pylori.

Bacilysin from Bacillus amyloliquefaciens FZB42 Has Specific Bactericidal Activity against Harmful Algal Bloom Species

PubMed Central

Wu, Liming; Wu, Huijun; Chen, Lina; Xie, Shanshan; Zang, Haoyu; Borriss, Rainer

2014-01-01

Harmful algal blooms, caused by massive and exceptional overgrowth of microalgae and cyanobacteria, are a serious environmental problem worldwide. In the present study, we looked for Bacillus strains with sufficiently strong anticyanobacterial activity to be used as biocontrol agents. Among 24 strains, Bacillus amyloliquefaciens FZB42 showed the strongest bactericidal activity against Microcystis aeruginosa, with a kill rate of 98.78%. The synthesis of the anticyanobacterial substance did not depend on Sfp, an enzyme that catalyzes a necessary processing step in the nonribosomal synthesis of lipopeptides and polyketides, but was associated with the aro gene cluster that is involved in the synthesis of the sfp-independent antibiotic bacilysin. Disruption of bacB, the gene in the cluster responsible for synthesizing bacilysin, or supplementation with the antagonist N-acetylglucosamine abolished the inhibitory effect, but this was restored when bacilysin synthesis was complemented. Bacilysin caused apparent changes in the algal cell wall and cell organelle membranes, and this resulted in cell lysis. Meanwhile, there was downregulated expression of glmS, psbA1, mcyB, and ftsZ—genes involved in peptidoglycan synthesis, photosynthesis, microcystin synthesis, and cell division, respectively. In addition, bacilysin suppressed the growth of other harmful algal species. In summary, bacilysin produced by B. amyloliquefaciens FZB42 has anticyanobacterial activity and thus could be developed as a biocontrol agent to mitigate the effects of harmful algal blooms. PMID:25261512

Molecular mechanisms of inflammation and tissue injury after major trauma-is complement the "bad guy"?

PubMed Central

2011-01-01

Trauma represents the leading cause of death among young people in industrialized countries. Recent clinical and experimental studies have brought increasing evidence for activation of the innate immune system in contributing to the pathogenesis of trauma-induced sequelae and adverse outcome. As the "first line of defense", the complement system represents a potent effector arm of innate immunity, and has been implicated in mediating the early posttraumatic inflammatory response. Despite its generic beneficial functions, including pathogen elimination and immediate response to danger signals, complement activation may exert detrimental effects after trauma, in terms of mounting an "innocent bystander" attack on host tissue. Posttraumatic ischemia/reperfusion injuries represent the classic entity of complement-mediated tissue damage, adding to the "antigenic load" by exacerbation of local and systemic inflammation and release of toxic mediators. These pathophysiological sequelae have been shown to sustain the systemic inflammatory response syndrome after major trauma, and can ultimately contribute to remote organ injury and death. Numerous experimental models have been designed in recent years with the aim of mimicking the inflammatory reaction after trauma and to allow the testing of new pharmacological approaches, including the emergent concept of site-targeted complement inhibition. The present review provides an overview on the current understanding of the cellular and molecular mechanisms of complement activation after major trauma, with an emphasis of emerging therapeutic concepts which may provide the rationale for a "bench-to-bedside" approach in the design of future pharmacological strategies. PMID:22129197

Streptococcus pyogenes Endopeptidase O Contributes to Evasion from Complement-mediated Bacteriolysis via Binding to Human Complement Factor C1q*

PubMed Central

Honda-Ogawa, Mariko; Sumitomo, Tomoko; Mori, Yasushi; Hamd, Dalia Talat; Ogawa, Taiji; Yamaguchi, Masaya; Nakata, Masanobu; Kawabata, Shigetada

2017-01-01

Streptococcus pyogenes secretes various virulence factors for evasion from complement-mediated bacteriolysis. However, full understanding of the molecules possessed by this organism that interact with complement C1q, an initiator of the classical complement pathway, remains elusive. In this study, we identified an endopeptidase of S. pyogenes, PepO, as an interacting molecule, and investigated its effects on complement immunity and pathogenesis. Enzyme-linked immunosorbent assay and surface plasmon resonance analysis findings revealed that S. pyogenes recombinant PepO bound to human C1q in a concentration-dependent manner under physiological conditions. Sites of inflammation are known to have decreased pH levels, thus the effects of PepO on bacterial evasion from complement immunity was analyzed in a low pH condition. Notably, under low pH conditions, PepO exhibited a higher affinity for C1q as compared with IgG, and PepO inhibited the binding of IgG to C1q. In addition, pepO deletion rendered S. pyogenes more susceptible to the bacteriocidal activity of human serum. Also, observations of the morphological features of the pepO mutant strain (ΔpepO) showed damaged irregular surfaces as compared with the wild-type strain (WT). WT-infected tissues exhibited greater severity and lower complement activity as compared with those infected by ΔpepO in a mouse skin infection model. Furthermore, WT infection resulted in a larger accumulation of C1q than that with ΔpepO. Our results suggest that interaction of S. pyogenes PepO with C1q interferes with the complement pathway, which enables S. pyogenes to evade complement-mediated bacteriolysis under acidic conditions, such as seen in inflammatory sites. PMID:28154192

Innate Immune Mechanisms in Transplant Allograft Vasculopathy

PubMed Central

Jane-wit, D; Fang, C; Goldstein, DR

2016-01-01

Purpose of Review Allograft vasculopathy (AV) is the leading cause of late allograft loss following solid organ transplantation. Ischemia reperfusion injury (IRI) and donor specific antibody (DSA)-induced complement activation confer heightened risk for AV via numerous innate immune mechanisms including MyD88, HMGB1, and complement induced non-canonical NF-kB signaling. Recent Findings The role of MyD88, a signal adaptor downstream of the toll-like receptors (TLR), has been defined in an experimental heart transplant model, which demonstrated that recipient MyD88 enhanced AV. Importantly, triggering receptor on myeloid receptor 1(Trem1), a MyD88 amplifying signal, was present in rejecting human cardiac transplant biopsies and enhanced the development of AV in mice. HMGB1, a nuclear protein that activates TLRs, also enhanced the development of AV. Complement activation elicits assembly of membrane attack complexes (MAC) on endothelial cells which activate non-canonical NF-kB signaling, a novel complement effector pathway that induces pro-inflammatory genes and potentiates endothelial cell mediated alloimmune T cell activation, processes which enhance AV. Summary Innate immune mediators including HMGB1, MyD88, and non-canonical NFκB signaling via complement activation contribute to AV. These pathways represent potential therapeutic targets to reduce AV after solid organ transplantation. PMID:27077602

Synergistic effect of a recombinant N-terminal fragment of bactericidal/permeability-increasing protein and cefamandole in treatment of rabbit gram-negative sepsis.

PubMed

Lin, Y; Leach, W J; Ammons, W S

1996-01-01

As a consequence of their bactericidal actions, many antibiotics cause the release of endotoxin, a primary mediator of gram-negative sepsis. Bactericidal/permeability-increasing protein (BPI) has bactericidal activity and neutralizes endotoxin in vitro and in vivo. We sought to examine the effect of a recombinant N-terminal fragment of BPI (rBPI21) in conjunction with cefamandole, a cephalosporin antibiotic, in the treatment of Escherichia coli bacteremia and septic shock in rabbits. Cefamandole (100 mg/kg of body weight) was injected intravenously. This was followed by simultaneous 10-min infusions of E. coli O7:K1 (9 x 10(9) CFU/kg) and rBPI21 (10 mg/kg). rBPI21 was continuously infused for an additional 110 min at 10 mg/kg/h. The administration of rBPI21 in conjunction with the administration of cefamandole prevented the cefamandole-induced increase of free endotoxin in plasma, accelerated bacterial clearance, ameliorated cardiopulmonary dysfunction, and thereby, prevented death, whereas neither agent alone was protective in this animal model. The efficacy of the combined treatment with rBPI21 and cefamandole suggests a synergistic interaction between the two agents. The data indicate that rBPI21 may be useful in conjunction with traditional antibiotic therapy.

Urinary tract infection: recent insight into the evolutionary arms race between uropathogenic Escherichia coli and our immune system.

PubMed

Schwab, Sebastian; Jobin, Katarzyna; Kurts, Christian

2017-12-01

Urinary tract infections (UTIs) are among the most common bacterial infections worldwide. Humans evolved various immune-dependent and independent defense mechanisms, while pathogens evolved multiple virulence factors to fight back. This article summarizes recent findings regarding the arms race between hosts and pathogens in UTIs. It was recently reported that macrophage subsets regulate neutrophil-mediated defense in primary UTIs but seem to subvert adaptive immunity upon re-infection. Moreover, some bacterial strains can survive inside macrophages, leading to recurrent infections. Inflammasome activation results in infected host cell death and pathogen release, facilitating the removal of intracellular bacteria. As a counteraction, some bacteria evolved mechanisms to disrupt inflammasome activation. Mucosal-associated invariant T cells are further effectors that can lyse infected epithelial cells and release intracellular bacteria. Once released, the bacteria are phagocytosed by neutrophils. However, some bacteria can inhibit neutrophil migration and deprive neutrophils of nutrients. Furthermore, the complement system, considered generally bactericidal, is exploited by the bacteria for cellular invasion. Another weapon against UTI is antimicrobial peptides, e.g. ribonuclease 7, but its production is inhibited by certain bacterial strains. Thus the arms race in UTI is ongoing, and knowing the enemy's methods can help in developing new drugs to win the race. © The Author 2017. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved.

Antimicrobial-induced endotoxin and cytokine activity in an in vitro model of septicemia in foals.

PubMed

Bentley, Adrienne P; Barton, Michelle H; Lee, Margie D; Norton, Natalie A; Moore, James N

2002-05-01

To determine which antimicrobials that are used to treat neonatal foals with septicemia attributable to Escherichia coli will minimize endotoxin release from bacteria and subsequent activity of inflammatory mediators while maintaining bactericidal efficacy. Blood samples from 10 healthy foals. Escherichia coli isolates A and B were isolated from 2 septicemic foals, and minimal inhibitory concentrations (MIC) were determined for 9 antimicrobials. Five of these antimicrobials were tested in vitro at 2 and 20 times their respective MIC. Whole blood or mononuclear cells grown in tissue-culture media were incubated with 105 colony-forming units of E. coli and each antimicrobial or saline (0.9% NaCl) solution. After 6 hours, number of viable bacteria remaining was determined, and supernatant was tested for endotoxin and tumor necrosis activity. Testing in whole blood was compromised by bactericidal effects of the blood itself. In mononuclear cell suspensions, each antimicrobial significantly reduced the number of viable bacteria to low or undetectable amounts. Antimicrobials did not differ significantly in efficacy of bacterial killing. Amikacin used alone or in combination with ampicillin resulted in significantly less endotoxin activity than did ampicillin, imipenem, or ceftiofur alone. There was a correlation between TNF-alpha and endotoxin activity. Aminoglycosides appear less likely to induce endotoxemia and TNF-alpha synthesis during bactericidal treatment of E. coli septicemia, compared with beta-lactam antimicrobials. Use of ampicillin, imipenem, or ceftiofur in the treatment of septicemic neonatal foals should be accompanied by appropriate treatment for endotoxemia.

The Lectin Complement Pathway Is Involved in Protection Against Enteroaggregative Escherichia coli Infection.

PubMed

Adler Sørensen, Camilla; Rosbjerg, Anne; Hebbelstrup Jensen, Betina; Krogfelt, Karen Angeliki; Garred, Peter

2018-01-01

Enteroaggregative Escherichia coli (EAEC) causes acute and persistent diarrhea worldwide. Still, the involvement of host factors in EAEC infections is unresolved. Binding of recognition molecules from the lectin pathway of complement to EAEC strains have been observed, but the importance is not known. Our aim was to uncover the involvement of these molecules in innate complement dependent immune protection toward EAEC. Binding of mannose-binding lectin, ficolin-1, -2, and -3 to four prototypic EAEC strains, and ficolin-2 binding to 56 clinical EAEC isolates were screened by a consumption-based ELISA method. Flow cytometry was used to determine deposition of C4b, C3b, and the bactericidal C5b-9 membrane attack complex (MAC) on the bacteria in combination with different complement inhibitors. In addition, the direct serum bactericidal effect was assessed. Screening of the prototypic EAEC strains revealed that ficolin-2 was the major binder among the lectin pathway recognition molecules. However, among the clinical EAEC isolates only a restricted number ( n  = 5) of the isolates bound ficolin-2. Using the ficolin-2 binding isolate C322-17 as a model, we found that incubation with normal human serum led to deposition of C4b, C3b, and to MAC formation. No inhibition of complement deposition was observed when a C1q inhibitor was added, while partial inhibition was observed when ficolin-2 or factor D inhibitors were used separately. Combining the inhibitors against ficolin-2 and factor D led to virtually complete inhibition of complement deposition and protection against direct bacterial killing. These results demonstrate that ficolin-2 may play an important role in innate immune protection against EAEC when an appropriate ligand is exposed, but many EAEC strains evade lectin pathway recognition and may, therefore, circumvent this strategy of innate host immune protection.

Exploring the Innate Immune System: Using Complement-Medicated Cell Lysis in the Classroom

ERIC Educational Resources Information Center

Fuller, Kevin G.

2008-01-01

The protein complement pathway comprises an important part of the innate immunity. The use of serum to demonstrate complement-mediated destruction across a series of bacterial dilutions allows an instructor to introduce a number of important biological concepts such as bacterial growth, activation cascades, and adaptive versus innate immunity.

The Human Antimicrobial Protein Bactericidal/Permeability-Increasing Protein (BPI) Inhibits the Infectivity of Influenza A Virus

PubMed Central

Pinkenburg, Olaf; Meyer, Torben; Bannert, Norbert; Norley, Steven; Bolte, Kathrin; Czudai-Matwich, Volker; Herold, Susanne; Gessner, André; Schnare, Markus

2016-01-01

In addition to their well-known antibacterial activity some antimicrobial peptides and proteins (AMPs) display also antiviral effects. A 27 aa peptide from the N-terminal part of human bactericidal/permeability-increasing protein (BPI) previously shown to harbour antibacterial activity inhibits the infectivity of multiple Influenza A virus strains (H1N1, H3N2 and H5N1) the causing agent of the Influenza pneumonia. In contrast, the homologous murine BPI-peptide did not show activity against Influenza A virus. In addition human BPI-peptide inhibits the activation of immune cells mediated by Influenza A virus. By changing the human BPI-peptide to the sequence of the mouse homologous peptide the antiviral activity was completely abolished. Furthermore, the human BPI-peptide also inhibited the pathogenicity of the Vesicular Stomatitis Virus but failed to interfere with HIV and measles virus. Electron microscopy indicate that the human BPI-peptide interferes with the virus envelope and at high concentrations was able to destroy the particles completely. PMID:27273104

Cluster Analysis Identifies Distinct Pathogenetic Patterns in C3 Glomerulopathies/Immune Complex-Mediated Membranoproliferative GN.

PubMed

Iatropoulos, Paraskevas; Daina, Erica; Curreri, Manuela; Piras, Rossella; Valoti, Elisabetta; Mele, Caterina; Bresin, Elena; Gamba, Sara; Alberti, Marta; Breno, Matteo; Perna, Annalisa; Bettoni, Serena; Sabadini, Ettore; Murer, Luisa; Vivarelli, Marina; Noris, Marina; Remuzzi, Giuseppe

2018-01-01

Membranoproliferative GN (MPGN) was recently reclassified as alternative pathway complement-mediated C3 glomerulopathy (C3G) and immune complex-mediated membranoproliferative GN (IC-MPGN). However, genetic and acquired alternative pathway abnormalities are also observed in IC-MPGN. Here, we explored the presence of distinct disease entities characterized by specific pathophysiologic mechanisms. We performed unsupervised hierarchical clustering, a data-driven statistical approach, on histologic, genetic, and clinical data and data regarding serum/plasma complement parameters from 173 patients with C3G/IC-MPGN. This approach divided patients into four clusters, indicating the existence of four different pathogenetic patterns. Specifically, this analysis separated patients with fluid-phase complement activation (clusters 1-3) who had low serum C3 levels and a high prevalence of genetic and acquired alternative pathway abnormalities from patients with solid-phase complement activation (cluster 4) who had normal or mildly altered serum C3, late disease onset, and poor renal survival. In patients with fluid-phase complement activation, those in clusters 1 and 2 had massive activation of the alternative pathway, including activation of the terminal pathway, and the highest prevalence of subendothelial deposits, but those in cluster 2 had additional activation of the classic pathway and the highest prevalence of nephrotic syndrome at disease onset. Patients in cluster 3 had prevalent activation of C3 convertase and highly electron-dense intramembranous deposits. In addition, we provide a simple algorithm to assign patients with C3G/IC-MPGN to specific clusters. These distinct clusters may facilitate clarification of disease etiology, improve risk assessment for ESRD, and pave the way for personalized treatment. Copyright © 2018 by the American Society of Nephrology.

C1q-Mediated Complement Activation and C3 Opsonization Trigger Recognition of Stealth Poly(2-methyl-2-oxazoline)-Coated Silica Nanoparticles by Human Phagocytes.

PubMed

Tavano, Regina; Gabrielli, Luca; Lubian, Elisa; Fedeli, Chiara; Visentin, Silvia; Polverino De Laureto, Patrizia; Arrigoni, Giorgio; Geffner-Smith, Alessandra; Chen, Fangfang; Simberg, Dmitri; Morgese, Giulia; Benetti, Edmondo M; Wu, Linping; Moghimi, Seyed Moein; Mancin, Fabrizio; Papini, Emanuele

2018-05-23

Poly(2-methyl-2-oxazoline) (PMOXA) is an alternative promising polymer to poly(ethylene glycol) (PEG) for design and engineering of macrophage-evading nanoparticles (NPs). Although PMOXA-engineered NPs have shown comparable pharmacokinetics and in vivo performance to PEGylated stealth NPs in the murine model, its interaction with elements of the human innate immune system has not been studied. From a translational angle, we studied the interaction of fully characterized PMOXA-coated vinyltriethoxysilane-derived organically modified silica NPs (PMOXA-coated NPs) of approximately 100 nm in diameter with human complement system, blood leukocytes, and macrophages and compared their performance with PEGylated and uncoated NP counterparts. Through detailed immunological and proteomic profiling, we show that PMOXA-coated NPs extensively trigger complement activation in human sera exclusively through the classical pathway. Complement activation is initiated by the sensing molecule C1q, where C1q binds with high affinity ( K d = 11 ± 1 nM) to NP surfaces independent of immunoglobulin binding. C1q-mediated complement activation accelerates PMOXA opsonization with the third complement protein (C3) through the amplification loop of the alternative pathway. This promoted NP recognition by human blood leukocytes and monocyte-derived macrophages. The macrophage capture of PMOXA-coated NPs correlates with sera donor variability in complement activation and opsonization but not with other major corona proteins, including clusterin and a wide range of apolipoproteins. In contrast to these observations, PMOXA-coated NPs poorly activated the murine complement system and were marginally recognized by mouse macrophages. These studies provide important insights into compatibility of engineered NPs with elements of the human innate immune system for translational steps.

Non-specific adsorption of complement proteins affects complement activation pathways of gold nanomaterials.

PubMed

Quach, Quang Huy; Kah, James Chen Yong

2017-04-01

The complement system is a key humoral component of innate immunity, serving as the first line of defense against intruders, including foreign synthetic nanomaterials. Although gold nanomaterials (AuNMs) are widely used in nanomedicine, their immunological response is not well understood. Using AuNMs of three shapes commonly used in biomedical applications: spherical gold nanoparticles, gold nanostars and gold nanorods, we demonstrated that AuNMs activated whole complement system, leading to the formation of SC5b-9 complex. All three complement pathways were simultaneously activated by all the AuNMs. Recognition molecules of the complement system interacted with all AuNMs in vitro, except for l-ficolin, but the correlation between these interactions and corresponding complement pathway activation was only observed in the classical and alternative pathways. We also observed the mediating role of complement activation in cellular uptake of all AuNMs by human U937 promonocytic cells, which expresses complement receptors. Taken together, our results highlighted the potential immunological challenges for clinical applications of AuNMs that were often overlooked.

Protection of host cells by complement regulators.

PubMed

Schmidt, Christoph Q; Lambris, John D; Ricklin, Daniel

2016-11-01

The complement cascade is an ancient immune-surveillance system that not only provides protection from pathogen invasion but has also evolved to participate in physiological processes to maintain tissue homeostasis. The alternative pathway (AP) of complement activation is the evolutionarily oldest part of this innate immune cascade. It is unique in that it is continuously activated at a low level and arbitrarily probes foreign, modified-self, and also unaltered self-structures. This indiscriminate activation necessitates the presence of preformed regulators on autologous surfaces to spare self-cells from the undirected nature of AP activation. Although the other two canonical complement activation routes, the classical and lectin pathways, initiate the cascade more specifically through pattern recognition, their activity still needs to be tightly controlled to avoid excessive reactivity. It is the perpetual duty of complement regulators to protect the self from damage inflicted by inadequate complement activation. Here, we review the role of complement regulators as preformed mediators of defense, explain their common and specialized functions, and discuss selected cases in which alterations in complement regulators lead to disease. Finally, rational engineering approaches using natural complement inhibitors as potential therapeutics are highlighted. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

A targeted complement-dependent strategy to improve the outcome of mAb therapy, and characterization in a murine model of metastatic cancer

PubMed Central

Elvington, Michelle; Huang, Yuxiang; Morgan, B. Paul; Qiao, Fei; van Rooijen, Nico; Atkinson, Carl

2012-01-01

Complement inhibitors expressed on tumor cells provide an evasion mechanism against mAb therapy and may modulate the development of an acquired antitumor immune response. Here we investigate a strategy to amplify mAb-targeted complement activation on a tumor cell, independent of a requirement to target and block complement inhibitor expression or function, which is difficult to achieve in vivo. We constructed a murine fusion protein, CR2Fc, and demonstrated that the protein targets to C3 activation products deposited on a tumor cell by a specific mAb, and amplifies mAb-dependent complement activation and tumor cell lysis in vitro. In syngeneic models of metastatic lymphoma (EL4) and melanoma (B16), CR2Fc significantly enhanced the outcome of mAb therapy. Subsequent studies using the EL4 model with various genetically modified mice and macrophage-depleted mice revealed that CR2Fc enhanced the therapeutic effect of mAb therapy via both macrophage-dependent FcγR-mediated antibody-dependent cellular cytotoxicity, and by direct complement-mediated lysis. Complement activation products can also modulate adaptive immunity, but we found no evidence that either mAb or CR2Fc treatment had any effect on an antitumor humoral or cellular immune response. CR2Fc represents a potential adjuvant treatment to increase the effectiveness of mAb therapy of cancer. PMID:22442351

Activity of Medicinal Plant Extracts on Multiplication of Mycobacterium tuberculosis under Reduced Oxygen Conditions Using Intracellular and Axenic Assays

PubMed Central

Bhatter, Purva D.; Gupta, Pooja D.; Birdi, Tannaz J.

2016-01-01

Aim. Test the activity of selected medicinal plant extracts on multiplication of Mycobacterium tuberculosis under reduced oxygen concentration which represents nonreplicating conditions. Material and Methods. Acetone, ethanol and aqueous extracts of the plants Acorus calamus L. (rhizome), Ocimum sanctum L. (leaf), Piper nigrum L. (seed), and Pueraria tuberosa DC. (tuber) were tested on Mycobacterium tuberculosis H37Rv intracellularly using an epithelial cell (A549) infection model. The extracts found to be active intracellularly were further studied axenically under reducing oxygen concentrations. Results and Conclusions. Intracellular multiplication was inhibited ≥60% by five of the twelve extracts. Amongst these 5 extracts, in axenic culture, P. nigrum (acetone) was active under aerobic, microaerophilic, and anaerobic conditions indicating presence of multiple components acting at different levels and P. tuberosa (aqueous) showed bactericidal activity under microaerophilic and anaerobic conditions implying the influence of anaerobiosis on its efficacy. P. nigrum (aqueous) and A. calamus (aqueous and ethanol) extracts were not active under axenic conditions but only inhibited intracellular growth of Mycobacterium tuberculosis, suggesting activation of host defense mechanisms to mediate bacterial killing rather than direct bactericidal activity. PMID:26941797

Human L-ficolin, a recognition molecule of the lectin activation pathway of complement, activates complement by binding to pneumolysin, the major toxin of Streptococcus pneumoniae.

PubMed

Ali, Youssif M; Kenawy, Hany I; Muhammad, Adnan; Sim, Robert B; Andrew, Peter W; Schwaeble, Wilhelm J

2013-01-01

The complement system is an essential component of the immune response, providing a critical line of defense against different pathogens including S. pneumoniae. Complement is activated via three distinct pathways: the classical (CP), the alternative (AP) and the lectin pathway (LP). The role of Pneumolysin (PLY), a bacterial toxin released by S. pneumoniae, in triggering complement activation has been studied in vitro. Our results demonstrate that in both human and mouse sera complement was activated via the CP, initiated by direct binding of even non-specific IgM and IgG3 to PLY. Absence of CP activity in C1q(-/-) mouse serum completely abolished any C3 deposition. However, C1q depleted human serum strongly opsonized PLY through abundant deposition of C3 activation products, indicating that the LP may have a vital role in activating the human complement system on PLY. We identified that human L-ficolin is the critical LP recognition molecule that drives LP activation on PLY, while all of the murine LP recognition components fail to bind and activate complement on PLY. This work elucidates the detailed interactions between PLY and complement and shows for the first time a specific role of the LP in PLY-mediated complement activation in human serum.

Human L-ficolin, a Recognition Molecule of the Lectin Activation Pathway of Complement, Activates Complement by Binding to Pneumolysin, the Major Toxin of Streptococcus pneumoniae

PubMed Central

Ali, Youssif M.; Kenawy, Hany I.; Muhammad, Adnan; Sim, Robert B.

2013-01-01

The complement system is an essential component of the immune response, providing a critical line of defense against different pathogens including S. pneumoniae. Complement is activated via three distinct pathways: the classical (CP), the alternative (AP) and the lectin pathway (LP). The role of Pneumolysin (PLY), a bacterial toxin released by S. pneumoniae, in triggering complement activation has been studied in vitro. Our results demonstrate that in both human and mouse sera complement was activated via the CP, initiated by direct binding of even non-specific IgM and IgG3 to PLY. Absence of CP activity in C1q−/− mouse serum completely abolished any C3 deposition. However, C1q depleted human serum strongly opsonized PLY through abundant deposition of C3 activation products, indicating that the LP may have a vital role in activating the human complement system on PLY. We identified that human L-ficolin is the critical LP recognition molecule that drives LP activation on PLY, while all of the murine LP recognition components fail to bind and activate complement on PLY. This work elucidates the detailed interactions between PLY and complement and shows for the first time a specific role of the LP in PLY-mediated complement activation in human serum. PMID:24349316

Indole-based novel small molecules for the modulation of bacterial signalling pathways.

PubMed

Biswas, Nripendra Nath; Kutty, Samuel K; Barraud, Nicolas; Iskander, George M; Griffith, Renate; Rice, Scott A; Willcox, Mark; Black, David StC; Kumar, Naresh

2015-01-21

Gram-negative bacteria such as Pseudomonas aeruginosa use N-acylated L-homoserine lactones (AHLs) as autoinducers (AIs) for quorum sensing (QS), a major regulatory and cell-to-cell communication system for social adaptation, virulence factor production, biofilm formation and antibiotic resistance. Some bacteria use indole moieties for intercellular signaling and as regulators of various bacterial phenotypes important for evading the innate host immune response and antimicrobial resistance. A range of natural and synthetic indole derivatives have been found to act as inhibitors of QS-dependent bacterial phenotypes, complementing the bactericidal ability of traditional antibiotics. In this work, various indole-based AHL mimics were designed and synthesized via the 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC·HCl) and N,N'-dicyclohexylcarbodiimide (DCC) mediated coupling reactions of a variety of substituted or unsubstituted aminoindoles with different alkanoic acids. All synthesized compounds were tested for QS inhibition using a P. aeruginosa QS reporter strain by measuring the amount of green fluorescent protein (GFP) production. Docking studies were performed to examine their potential to bind and therefore inhibit the target QS receptor protein. The most potent compounds 11a, 11d and 16a showed 44 to 65% inhibition of QS activity at 250 μM concentration, and represent promising drug leads for the further development of anti-QS antimicrobial compounds.

Complement-Mediated Regulation of Metabolism and Basic Cellular Processes.

PubMed

Hess, Christoph; Kemper, Claudia

2016-08-16

Complement is well appreciated as a critical arm of innate immunity. It is required for the removal of invading pathogens and works by directly destroying them through the activation of innate and adaptive immune cells. However, complement activation and function is not confined to the extracellular space but also occurs within cells. Recent work indicates that complement activation regulates key metabolic pathways and thus can impact fundamental cellular processes, such as survival, proliferation, and autophagy. Newly identified functions of complement include a key role in shaping metabolic reprogramming, which underlies T cell effector differentiation, and a role as a nexus for interactions with other effector systems, in particular the inflammasome and Notch transcription-factor networks. This review focuses on the contributions of complement to basic processes of the cell, in particular the integration of complement with cellular metabolism and the potential implications in infection and other disease settings. Copyright © 2016 Elsevier Inc. All rights reserved.

Complement system and immunological mediators: Their involvements in the induced inflammatory process by Androctonus australis hector venom and its toxic components.

PubMed

Bekkari, Nadjia; Martin-Eauclaire, Marie-France; Laraba-Djebari, Fatima

2015-01-01

Androctonus australis hector scorpion venom is well known by its high toxicity, it induces massive release of neurotransmitters that lead to pathophysiological disorders in cardiovascular, neuro-hormonal and immune systems. Previous studies have shown the relationship between the severity of scorpion envenoming and immune system activation. This study was assessed to investigate the involvement of complement system and inflammatory mediators after sublethal injection of Aah venom, its toxic fraction (FtoxG50) and its main toxins (AahI and AahII) into NMRI mice. The Activation complement system by the venom is also compared to that induced of lipopolysaccharides (LPS). Obtained results showed that seric complement system (CS) is activated by the venom and by its toxic components; this activation is more pronounced into liver tissue when toxic components (FtoxG50, AahI or AahII) are used. Increase of cytokine levels (IL1β, TNFα and ICAM) into hepatic tissue induced by AahI or AahII neurotoxins is correlated with tissue alterations. Aprotinin, a non specific inhibitor of complement system seems to be able to reduce CS consumption and to restore partially the induced tissue damage by venom. The mechanisms by which toxic fraction or LPS induced the activation of complement system seem to be different. Sensitivity of hepatic tissue is more pronounced after FtoxG50 injection; however lung tissue is more sensible to LPS than FoxG50. Copyright © 2015 Elsevier GmbH. All rights reserved.

THE BACTERICIDAL ACTIVITY OF NORMAL GUINEA PIG SERUM AGAINST LISTERIA MONOCYTOGENES AND ITS INHIBITION BY A LISTERIAL CELL EXTRACT,

DTIC Science & Technology

Normal guinea pig serum contains bactericidins active against Listeria monocytogenes. The listeriocidal activity of the serum did not increase after...factor. Lysozyme was not implicated in the bactericidal system. It was suggested that the bactericidal activity of guinea pig serum might be due either to

Complement Activation in Arterial and Venous Thrombosis is Mediated by Plasmin

PubMed Central

Foley, Jonathan H.; Walton, Bethany L.; Aleman, Maria M.; O'Byrne, Alice M.; Lei, Victor; Harrasser, Micaela; Foley, Kimberley A.; Wolberg, Alisa S.; Conway, Edward M.

2016-01-01

Thrombus formation leading to vaso-occlusive events is a major cause of death, and involves complex interactions between coagulation, fibrinolytic and innate immune systems. Leukocyte recruitment is a key step, mediated partly by chemotactic complement activation factors C3a and C5a. However, mechanisms mediating C3a/C5a generation during thrombosis have not been studied. In a murine venous thrombosis model, levels of thrombin–antithrombin complexes poorly correlated with C3a and C5a, excluding a central role for thrombin in C3a/C5a production. However, clot weight strongly correlated with C5a, suggesting processes triggered during thrombosis promote C5a generation. Since thrombosis elicits fibrinolysis, we hypothesized that plasmin activates C5 during thrombosis. In vitro, the catalytic efficiency of plasmin-mediated C5a generation greatly exceeded that of thrombin or factor Xa, but was similar to the recognized complement C5 convertases. Plasmin-activated C5 yielded a functional membrane attack complex (MAC). In an arterial thrombosis model, plasminogen activator administration increased C5a levels. Overall, these findings suggest plasmin bridges thrombosis and the immune response by liberating C5a and inducing MAC assembly. These new insights may lead to the development of strategies to limit thrombus formation and/or enhance resolution. PMID:27077125

Identification of a defined linear epitope in the OspA protein of the Lyme disease spirochetes that elicits bactericidal antibody responses: Implications for vaccine development.

PubMed

Izac, Jerilyn R; Oliver, Lee D; Earnhart, Christopher G; Marconi, Richard T

2017-05-31

The lipoprotein OspA is produced by the Lyme disease spirochetes primarily in unfed ticks. OspA production is down-regulated by the blood meal and it is not produced in mammals except for possible transient production during late stage infection in patients with Lyme arthritis. Vaccination with OspA elicits antibody (Ab) that can target spirochetes in the tick midgut during feeding and inhibit transmission to mammals. OspA was the primary component of the human LYMErix™ vaccine. LYMErix™ was available from 1998 to 2002 but then pulled from the market due to declining sales as a result of unsubstantiated concerns about vaccination induced adverse events and poor efficacy. It was postulated that a segment of OspA that shares sequence similarity with a region in human LFA-1 and may trigger putative autoimmune events. While evidence supporting such a link has not been demonstrated, most efforts to move forward with OspA as a vaccine component have sought to eliminate this region of concern. Here we identify an OspA linear epitope localized within OspA amino acid residues 221-240 (OspA 221-240 ) that lacks the OspA region suggested to elicit autoimmunity. A peptide consisting of residues 221-240 was immunogenic in mice. Ab raised against OspA 221-240 peptide surface labeled B. burgdorferi in IFAs and displayed potent Ab mediated-complement dependent bactericidal activity. BLAST analyses identified several variants of OspA 221-240 and a closely related sequence in OspB. It is our hypothesis that integration of the OspA 221-240 epitope into a multivalent-OspC based chimeric epitope based vaccine antigen (chimeritope) could result in a subunit vaccine that protects against Lyme disease through synergistic mechanisms. Copyright © 2017. Published by Elsevier Ltd.

Bactericidal activity of antibiotics against Legionella micdadei (Pittsburgh pneumonia agent).

PubMed Central

Dowling, J N; Weyant, R S; Pasculle, A W

1982-01-01

The bactericidal activity of five antibiotics for Legionella micdadei was determined by the construction of time-kill curves. Erythromycin, rifampin, penicillin G, cephalothin, and gentamicin were bactericidal for L. micdadei at readily achievable concentrations. The minimal bactericidal concentrations, defined as those producing 99.9% killing within 24 h, were: erythromycin, 4.6; rifampin, 0.13; penicillin G, 0.25; cephalothin, 2.5; and gentamicin, 0.25 micrograms/ml. The ratios of the minimal bactericidal to minimal inhibitory concentrations for these antibiotics ranged from 1 to 8. Thus, the poor in vivo activity of beta-lactam and aminoglycoside antibiotics against L. micdadei cannot be ascribed to a lack of killing by these agents. PMID:6927637

Streptococcus pyogenes Endopeptidase O Contributes to Evasion from Complement-mediated Bacteriolysis via Binding to Human Complement Factor C1q.

PubMed

Honda-Ogawa, Mariko; Sumitomo, Tomoko; Mori, Yasushi; Hamd, Dalia Talat; Ogawa, Taiji; Yamaguchi, Masaya; Nakata, Masanobu; Kawabata, Shigetada

2017-03-10

Streptococcus pyogenes secretes various virulence factors for evasion from complement-mediated bacteriolysis. However, full understanding of the molecules possessed by this organism that interact with complement C1q, an initiator of the classical complement pathway, remains elusive. In this study, we identified an endopeptidase of S. pyogenes , PepO, as an interacting molecule, and investigated its effects on complement immunity and pathogenesis. Enzyme-linked immunosorbent assay and surface plasmon resonance analysis findings revealed that S. pyogenes recombinant PepO bound to human C1q in a concentration-dependent manner under physiological conditions. Sites of inflammation are known to have decreased pH levels, thus the effects of PepO on bacterial evasion from complement immunity was analyzed in a low pH condition. Notably, under low pH conditions, PepO exhibited a higher affinity for C1q as compared with IgG, and PepO inhibited the binding of IgG to C1q. In addition, pepO deletion rendered S. pyogenes more susceptible to the bacteriocidal activity of human serum. Also, observations of the morphological features of the pepO mutant strain (Δ pepO ) showed damaged irregular surfaces as compared with the wild-type strain (WT). WT-infected tissues exhibited greater severity and lower complement activity as compared with those infected by Δ pepO in a mouse skin infection model. Furthermore, WT infection resulted in a larger accumulation of C1q than that with Δ pepO. Our results suggest that interaction of S. pyogenes PepO with C1q interferes with the complement pathway, which enables S. pyogenes to evade complement-mediated bacteriolysis under acidic conditions, such as seen in inflammatory sites. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

The Siderophore Product Ornibactin Is Required for the Bactericidal Activity of Burkholderia contaminans MS14.

PubMed

Deng, Peng; Foxfire, Adam; Xu, Jianhong; Baird, Sonya M; Jia, Jiayuan; Delgado, Keren H; Shin, Ronald; Smith, Leif; Lu, Shi-En

2017-04-15

Burkholderia contaminans MS14 was isolated from soil in Mississippi. When it is cultivated on nutrient broth-yeast extract agar, the colonies exhibit bactericidal activity against a wide range of plant-pathogenic bacteria. A bacteriostatic compound with siderophore activity was successfully purified and was determined by nuclear magnetic resonance spectroscopy to be ornibactin. Isolation of the bactericidal compound has not yet been achieved; therefore, the exact nature of the bactericidal compound is still unknown. During an attempt to isolate the bactericidal compound, an interesting relationship between the production of ornibactin and the bactericidal activity of MS14 was characterized. Transposon mutagenesis resulted in two strains that lost bactericidal activity, with insertional mutations in a nonribosomal peptide synthetase (NRPS) gene for ornibactin biosynthesis and a luxR family transcriptional regulatory gene. Coculture of these two mutant strains resulted in restoration of the bactericidal activity. Furthermore, the addition of ornibactin to the NRPS mutant restored the bactericidal phenotype. It has been demonstrated that, in MS14, ornibactin has an alternative function, aside from iron sequestration. Comparison of the ornibactin biosynthesis genes in Burkholderia species shows diversity among the regulatory elements, while the gene products for ornibactin synthesis are conserved. This is an interesting observation, given that ornibactin is thought to have the same defined function within Burkholderia species. Ornibactin is produced by most Burkholderia species, and its role in regulating the production of secondary metabolites should be investigated. IMPORTANCE Identification of the antibacterial product from strain MS14 is not the key feature of this study. We present a series of experiments that demonstrate that ornibactin is directly involved in the bactericidal phenotype of MS14. This observation provides evidence for an alternative function for ornibactin, aside from iron sequestration. Ornibactin should be further evaluated for its role in regulating the biosynthesis of secondary metabolites in other Burkholderia species. Copyright © 2017 American Society for Microbiology.

The Siderophore Product Ornibactin Is Required for the Bactericidal Activity of Burkholderia contaminans MS14

PubMed Central

Deng, Peng; Foxfire, Adam; Xu, Jianhong; Baird, Sonya M.; Jia, Jiayuan; Delgado, Keren H.; Shin, Ronald

2017-01-01

ABSTRACT Burkholderia contaminans MS14 was isolated from soil in Mississippi. When it is cultivated on nutrient broth-yeast extract agar, the colonies exhibit bactericidal activity against a wide range of plant-pathogenic bacteria. A bacteriostatic compound with siderophore activity was successfully purified and was determined by nuclear magnetic resonance spectroscopy to be ornibactin. Isolation of the bactericidal compound has not yet been achieved; therefore, the exact nature of the bactericidal compound is still unknown. During an attempt to isolate the bactericidal compound, an interesting relationship between the production of ornibactin and the bactericidal activity of MS14 was characterized. Transposon mutagenesis resulted in two strains that lost bactericidal activity, with insertional mutations in a nonribosomal peptide synthetase (NRPS) gene for ornibactin biosynthesis and a luxR family transcriptional regulatory gene. Coculture of these two mutant strains resulted in restoration of the bactericidal activity. Furthermore, the addition of ornibactin to the NRPS mutant restored the bactericidal phenotype. It has been demonstrated that, in MS14, ornibactin has an alternative function, aside from iron sequestration. Comparison of the ornibactin biosynthesis genes in Burkholderia species shows diversity among the regulatory elements, while the gene products for ornibactin synthesis are conserved. This is an interesting observation, given that ornibactin is thought to have the same defined function within Burkholderia species. Ornibactin is produced by most Burkholderia species, and its role in regulating the production of secondary metabolites should be investigated. IMPORTANCE Identification of the antibacterial product from strain MS14 is not the key feature of this study. We present a series of experiments that demonstrate that ornibactin is directly involved in the bactericidal phenotype of MS14. This observation provides evidence for an alternative function for ornibactin, aside from iron sequestration. Ornibactin should be further evaluated for its role in regulating the biosynthesis of secondary metabolites in other Burkholderia species. PMID:28188204

[Bactericidal activity of serum and chemotherapy in sensitive and resistant exciter (author's transl)].

PubMed

Eyer, H; Metz, H; Preac-Mursic, V

1975-11-21

Comparing examinations with Ampicillin sensitive and resistant bacteria-strains show that the bactericidal activity of serum is dependent on the bacteria-strains, on the Ampicillin sensitivity of the particular exciter and on the number of bacteria/ml (germ count). Bactericide effect could always be obtained with sensitive strains as a result of additional chemotherapy. With several resistant strains a bactericide effect could not be obtained in this case the continuous optimal Ampicillin addition was the decisive factor. Because of the extremely complicated process of the bactericide one should not make general conclusions from the individual experimental results.

Classical and alternative complement activation on photoreceptor outer segments drives monocyte-dependent retinal atrophy.

PubMed

Katschke, Kenneth J; Xi, Hongkang; Cox, Christian; Truong, Tom; Malato, Yann; Lee, Wyne P; McKenzie, Brent; Arceo, Rommel; Tao, Jianhua; Rangell, Linda; Reichelt, Mike; Diehl, Lauri; Elstrott, Justin; Weimer, Robby M; Campagne, Menno van Lookeren

2018-05-09

Geographic atrophy (GA), the advanced form of dry age-related macular degeneration (AMD), is characterized by progressive loss of retinal pigment epithelium cells and photoreceptors in the setting of characteristic extracellular deposits and remains a serious unmet medical need. While genetic predisposition to AMD is dominated by polymorphisms in complement genes, it remains unclear how complement activation contributes to retinal atrophy. Here we demonstrate that complement is activated on photoreceptor outer segments (POS) in the retina peripheral to atrophic lesions associated with GA. When exposed to human serum following outer blood-retinal barrier breakdown, POS act as potent activators of the classical and alternative complement pathway. In mouse models of retinal degeneration, classical and alternative pathway complement activation on photoreceptors contributed to the loss of photoreceptor function. This was dependent on C5a-mediated recruitment of peripheral blood monocytes but independent of resident microglia. Genetic or pharmacologic inhibition of both classical and alternative complement C3 and C5 convertases was required to reduce progressive degeneration of photoreceptor rods and cones. Our study implicates systemic classical and alternative complement proteins and peripheral blood monocytes as critical effectors of localized retinal degeneration with potential relevance for the contribution of complement activation to GA.

In vitro bacterial cytotoxicity of CNTs: reactive oxygen species mediate cell damage edges over direct physical puncturing.

PubMed

Rajavel, Krishnamoorthy; Gomathi, Rajkumar; Manian, Sellamuthu; Rajendra Kumar, Ramasamy Thangavelu

2014-01-21

Understanding the bacterial cytotoxicity of CNTs is important for a wide variety of applications in the biomedical, environmental, and health sectors. A majority of the earlier reports attributed the bactericidal cytotoxicity of CNTs to bacterial cell membrane damage by direct physical puncturing. Our results reveal that bacterial cell death via bacterial cell membrane damage is induced by reactive oxygen species (ROS) produced from CNTs and is not due to direct physical puncturing by CNTs. To understand the actual mechanism of bacterial killing, we elucidated the bacterial cytotoxicity of SWCNTs and MWCNTs against Gram-negative human pathogenic bacterial species Escherichia coli, Shigella sonnei, Klebsiella pneumoniae, and Pseudomonas aeruginosa and its amelioration upon functionalizing the CNTs with antioxidant tannic acid (TA). Interestingly, the bacterial cells treated with CNTs exhibited severe cell damage under laboratory (ambient) and sunlight irradiation conditions. However, CNTs showed no cytotoxicity to the bacterial cells when incubated in the dark. The quantitative assessments carried out by us made it explicit that CNTs are effective generators of ROS such as (1)O2, O2(•-), and (•)OH in an aqueous medium under both ambient and sunlight-irradiated conditions. Both naked and TA-functionalized CNTs showed negligible ROS production in the dark. Furthermore, strong correlations were obtained between ROS produced by CNTs and the bacterial cell mortality (with the correlation coefficient varying between 0.7618 and 0.9891) for all four tested pathogens. The absence of bactericidal cytotoxicity in both naked and functionalized CNTs in the dark reveals that the presence of ROS is the major factor responsible for the bactericidal action compared to direct physical puncturing. This understanding of the bactericidal activity of the irradiated CNTs, mediated through the generation of ROS, could be interesting for novel applications such as regulated ROS delivery in cancer therapy and the sanitation of potable water supplies.

Human milk bactericidal properties: effect of lyophilization and relation to maternal factors and milk components.

PubMed

Salcedo, Jaime; Gormaz, Maria; López-Mendoza, Maria C; Nogarotto, Elisabetta; Silvestre, Dolores

2015-04-01

Lyophilization appears to be a viable method for storing human milk, assuring no microbiological contamination and preserving its health benefits and antibacterial properties. The aim of the study is to evaluate and compare the effects of different storage methods (lyophilization and freezing at -20°C and -80°C) and maternal factors (gestational length or time postpartum) upon the microbiological contents and bactericidal activity of human milk. The possible relation between bactericidal activity and the content of certain nutrients and functional components is also investigated. Microbiological content, bactericidal activity, sialic acid, and ganglioside contents, as well as protein, fat, and lactose concentrations were assessed in 125 human milk samples from 65 healthy donors in the Human Milk Bank of La Fe (Valencia, Spain). Lyophilization and storage at -80°C significantly reduced the content of mesophilic aerobic microorganisms and Staphylococcus epidermidis when compared with storage at -20°C. Bactericidal activity was not significantly modified by lyophilization when compared with freezing at either -20°C or -80°C. Bactericidal activity was not correlated with fat, protein, or lactose content, but was significantly correlated to ganglioside content. The bactericidal activity was significantly greater (P < 0.05) in mature milk and in milk from women with term delivery than in milk from early lactation (days 1-7 postpartum) and milk from women with preterm delivery, respectively. Lyophilization and storage at -80°C of human milk yields similar results and are superior to storage at -20C with regard to microbial and bactericidal capacities, being a feasible alternative for human milk banks.

Engineered Fc variant antibodies with enhanced ability to recruit complement and mediate effector functions

PubMed Central

Moore, Gregory L; Chen, Hsing; Karki, Sher

2010-01-01

Engineering the antibody Fc region to enhance the cytotoxic activity of therapeutic antibodies is currently an active area of investigation. The contribution of complement to the mechanism of action of some antibodies that target cancers and pathogens makes a compelling case for its optimization. Here we describe the generation of a series of Fc variants with enhanced ability to recruit complement. Variants enhanced the cytotoxic potency of an anti-CD20 antibody up to 23-fold against tumor cells in CDC assays, and demonstrated a correlated increase in C1q binding affinity. Complementenhancing substitutions combined additively, and in one case synergistically, with substitutions previously engineered for improved binding to Fc gamma receptors. The engineered combinations provided a range of effector function activities, including simultaneously enhanced CDC, ADCC, and phagocytosis. Variants were also effective at boosting the effector function of antibodies targeting the antigens CD40 and CD19, in the former case enhancing CDC over 600-fold, and in the latter case imparting complement-mediated activity onto an IgG1 antibody that was otherwise incapable of it. This work expands the toolkit of modifications for generating monoclonal antibodies with improved therapeutic potential and enables the exploration of optimized synergy between Fc gamma receptors and complement pathways for the destruction of tumors and infectious pathogens. PMID:20150767

In Vitro Antibacterial Activities of AF 3013, the Active Metabolite of Prulifloxacin, against Nosocomial and Community Italian Isolates

PubMed Central

Montanari, Maria Pia; Mingoia, Marina; Varaldo, Pietro Emanuele

2001-01-01

AF 3013, the active metabolite of prulifloxacin, was tested to determine its inhibitory and bactericidal activities against 396 nosocomial and 258 community Italian isolates. Compared with that of ciprofloxacin, its activity (assessed in MIC and minimal bactericidal concentration tests) was generally similar or greater against gram-positive bacteria and greater against gram-negative bacteria. In time-kill assays using selected isolates, its bactericidal activity was comparable to that of ciprofloxacin. PMID:11709353

Bisphosphocins: novel antimicrobials for enhanced killing of drug-resistant and biofilm-forming bacteria.

PubMed

Wong, Jonathan P; DiTullio, Paul; Parkinson, Steve

2015-01-01

The global prevalence of antibiotic resistance and the threat posed by drug-resistant superbugs are a leading challenge confronting modern medicine in the 21st century. However, the progress on the development of novel antibiotics to combat this problem is severely lagging. A more concerted effort to develop novel therapeutic agents with robust activity and unique mechanisms of action will be needed to overcome the problem of drug resistance. Furthermore, biofilm forming bacteria are known to be increasingly resistant to the actions of antibiotics and are a leading cause of mortality or morbidity in nosocomial infections. Bisphosphocins (also scientifically known as nubiotics) are novel small protonated deoxynucleotide molecules, and exert their antibacterial activity by depolarization of the bacterial cell membrane, causing bacterial cell death. Bisphosphocins may represent an effective weapon against antibiotic-resistant and biofilm-forming pathogenic bacteria. Preclinical efficacy studies in animals have shown that the compounds are safe and, efficacious against various bacterial infections, including drug-resistant pathogens. In vitro biochemical analysis confirmed that the bactericidal activity of bisphosphocins is mediated by depolarization of the bacterial cell membrane, and these compounds are better able to penetrate through bacterial biofilm and kill the biofilm encased bacteria. This article will cover the structure, mode of action, safety, efficacy and the current state of development of bisphosphocins. Together, the information presented here will present a strong case for bisphosphocins to be considered for use as new weapons to complement the existing arsenal of antimicrobial drugs and as a first line defence against drug-resistant and biofilm-forming bacteria.

The important and diverse roles of antibodies in the host response to Borrelia infections.

PubMed

LaRocca, T J; Benach, J L

2008-01-01

Antibodies are of critical importance in the host response to tick-borne Borrelia species that cause relapsing fever and Lyme disease. Recent studies on the role of various B cell subsets in the host response to Borrelia, complement-independent, bactericidal antibodies, and diagnostics led to this review that focuses on the array of functions that antibodies to Borrelia can perform.

Mannose-binding lectin binds to Ebola and Marburg envelope glycoproteins, resulting in blocking of virus interaction with DC-SIGN and complement-mediated virus neutralization.

PubMed

Ji, Xin; Olinger, Gene G; Aris, Sheena; Chen, Ying; Gewurz, Henry; Spear, Gregory T

2005-09-01

Mannose-binding lectin (MBL), a serum lectin that mediates innate immune functions including activation of the lectin complement pathway, binds to carbohydrates expressed on some viral glycoproteins. In this study, the ability of MBL to bind to virus particles pseudotyped with Ebola and Marburg envelope glycoproteins was evaluated. Virus particles bearing either Ebola (Zaire strain) or Marburg (Musoke strain) envelope glycoproteins bound at significantly higher levels to immobilized MBL compared with virus particles pseudotyped with vesicular stomatitis virus glycoprotein or with no virus glycoprotein. As observed in previous studies, Ebola-pseudotyped virus bound to cells expressing the lectin DC-SIGN (dendritic cell-specific intercellular adhesion molecule 3-grabbing non-integrin). However, pre-incubation of virus with MBL blocked DC-SIGN-mediated binding to cells, suggesting that the two lectins bind at the same or overlapping sites on the Ebola glycoprotein. Neutralization experiments showed that virus pseudotyped with Ebola or Marburg (Musoke) glycoprotein was neutralized by complement, while the Marburg (Ravn strain) glycoprotein-pseudotyped virus was less sensitive to neutralization. Neutralization was partially mediated through the lectin complement pathway, since a complement source deficient in MBL was significantly less effective at neutralizing viruses pseudotyped with filovirus glycoproteins and addition of purified MBL to the MBL-deficient complement increased neutralization. These experiments demonstrated that MBL binds to filovirus envelope glycoproteins resulting in important biological effects and suggest that MBL can interact with filoviruses during infection in humans.

Mannose Binding Lectin Is Required for Alphavirus-Induced Arthritis/Myositis

PubMed Central

Whitmore, Alan C.; Blevins, Lance K.; Hueston, Linda; Fraser, Robert J.; Herrero, Lara J.; Ramirez, Ruben; Smith, Paul N.; Mahalingam, Suresh; Heise, Mark T.

2012-01-01

Mosquito-borne alphaviruses such as chikungunya virus and Ross River virus (RRV) are emerging pathogens capable of causing large-scale epidemics of virus-induced arthritis and myositis. The pathology of RRV-induced disease in both humans and mice is associated with induction of the host inflammatory response within the muscle and joints, and prior studies have demonstrated that the host complement system contributes to development of disease. In this study, we have used a mouse model of RRV-induced disease to identify and characterize which complement activation pathways mediate disease progression after infection, and we have identified the mannose binding lectin (MBL) pathway, but not the classical or alternative complement activation pathways, as essential for development of RRV-induced disease. MBL deposition was enhanced in RRV infected muscle tissue from wild type mice and RRV infected MBL deficient mice exhibited reduced disease, tissue damage, and complement deposition compared to wild-type mice. In contrast, mice deficient for key components of the classical or alternative complement activation pathways still developed severe RRV-induced disease. Further characterization of MBL deficient mice demonstrated that similar to C3−/− mice, viral replication and inflammatory cell recruitment were equivalent to wild type animals, suggesting that RRV-mediated induction of complement dependent immune pathology is largely MBL dependent. Consistent with these findings, human patients diagnosed with RRV disease had elevated serum MBL levels compared to healthy controls, and MBL levels in the serum and synovial fluid correlated with severity of disease. These findings demonstrate a role for MBL in promoting RRV-induced disease in both mice and humans and suggest that the MBL pathway of complement activation may be an effective target for therapeutic intervention for humans suffering from RRV-induced arthritis and myositis. PMID:22457620

Lactobacillus pentosus strain LPS16 produces lactic acid, inhibiting multidrug-resistant Helicobacter pylori.

PubMed

Zheng, Po-Xing; Fang, Hsin-Yi; Yang, Hsiao-Bai; Tien, Nai-Yueh; Wang, Ming-Cheng; Wu, Jiunn-Jong

2016-04-01

Helicobacter pylori is a human gastric pathogen. Antibiotic resistance of H. pylori has become a problem increasing the failure of H. pylori eradication. Therefore alternative approaches are required. The aim of this study was to evaluate the anti-H. pylori activity of Lactobacillus pentosus strain LPS16 and the mechanism of its killing effect. The anti-H. pylori activity of LPS16 was determined by the disc diffusion test and time killing assay. High-performance liquid chromatography analysis was used to analyze the secreted compounds of LPS16. Sixty H. pylori strains isolated from different gastric diseases, having different antibiotic susceptibility were collected to analyze the spectrum of anti-H. pylori activity of LPS16. Adhesion ability of LPS16 to gastric epithelial cell lines was assayed by flow cytometry. The anti-H. pylori activity of LPS16 depended on the secreted component, and lactic acid mediated bactericidal activity against H. pylori. The bactericidal activity did not vary significantly among the strains isolated from different diseases having different antibiotic susceptibility. Moreover, LPS16 can adhere on gastric epithelial cell lines AKG and MKN45. L. pentosus strain LPS16 had the broad-spectrum anti-H. pylori activity, suggesting that it can be used to prevent H. pylori infection. Copyright © 2014. Published by Elsevier B.V.

Serum bactericidal assay for the evaluation of typhoid vaccine using a semi-automated colony-counting method.

PubMed

Jang, Mi Seon; Sahastrabuddhe, Sushant; Yun, Cheol-Heui; Han, Seung Hyun; Yang, Jae Seung

2016-08-01

Typhoid fever, mainly caused by Salmonella enterica serovar Typhi (S. Typhi), is a life-threatening disease, mostly in developing countries. Enzyme-linked immunosorbent assay (ELISA) is widely used to quantify antibodies against S. Typhi in serum but does not provide information about functional antibody titers. Although the serum bactericidal assay (SBA) using an agar plate is often used to measure functional antibody titers against various bacterial pathogens in clinical specimens, it has rarely been used for typhoid vaccines because it is time-consuming and labor-intensive. In the present study, we established an improved SBA against S. Typhi using a semi-automated colony-counting system with a square agar plate harboring 24 samples. The semi-automated SBA efficiently measured bactericidal titers of sera from individuals immunized with S. Typhi Vi polysaccharide vaccines. The assay specifically responded to S. Typhi Ty2 but not to other irrelevant enteric bacteria including Vibrio cholerae and Shigella flexneri. Baby rabbit complement was more appropriate source for the SBA against S. Typhi than complements from adult rabbit, guinea pig, and human. We also examined the correlation between SBA and ELISA for measuring antibody responses against S. Typhi using pre- and post-vaccination sera from 18 human volunteers. The SBA titer showed a good correlation with anti-Vi IgG quantity in the serum as determined by Spearman correlation coefficient of 0.737 (P < 0.001). Taken together, the semi-automated SBA might be efficient, accurate, sensitive, and specific enough to measure functional antibody titers against S. Typhi in sera from human subjects immunized with typhoid vaccines. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Eotaxin-3 (CCL26) exerts innate host defense activities that are modulated by mast cell proteases.

PubMed

Gela, A; Kasetty, G; Jovic, S; Ekoff, M; Nilsson, G; Mörgelin, M; Kjellström, S; Pease, J E; Schmidtchen, A; Egesten, A

2015-02-01

During bacterial infections of the airways, a Th1-profiled inflammation promotes the production of several host defense proteins and peptides with antibacterial activities including β-defensins, ELR-negative CXC chemokines, and the cathelicidin LL-37. These are downregulated by Th2 cytokines of the allergic response. Instead, the eosinophil-recruiting chemokines eotaxin-1/CCL11, eotaxin-2/CCL24, and eotaxin-3/CCL26 are expressed. This study set out to investigate whether these chemokines could serve as innate host defense molecules during allergic inflammation. Antibacterial activities of the eotaxins were investigated using viable count assays, electron microscopy, and methods assessing bacterial permeabilization. Fragments generated by mast cell proteases were characterized, and their potential antibacterial, receptor-activating, and lipopolysaccharide-neutralizing activities were investigated. CCL11, CCL24, and CCL26 all showed potent bactericidal activity, mediated through membrane disruption, against the airway pathogens Streptococcus pneumoniae, Staphylococcus aureus, Nontypeable Haemophilus influenzae, and Pseudomonas aeruginosa. CCL26 retained bactericidal activity in the presence of salt at physiologic concentrations, and the region holding the highest bactericidal activity was the cationic and amphipathic COOH-terminus. Proteolysis of CCL26 by chymase and tryptase, respectively, released distinct fragments of the COOH- and NH2 -terminal regions. The COOH-terminal fragment retained antibacterial activity while the NH2 -terminal had potent LPS-neutralizing properties in the order of CCL26 full-length protein. An identical fragment to NH2 -terminal fragment generated by tryptase was obtained after incubation with supernatants from activated mast cells. None of the fragments activated the CCR3-receptor. Taken together, the findings show that the eotaxins can contribute to host defense against common airway pathogens and that their activities are modulated by mast cell proteases. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Autocrine Complement Inhibits IL10-Dependent T-Cell Mediated Antitumor Immunity to Promote Tumor Progression

PubMed Central

Wang, Yu; Sun, Sheng-Nan; Liu, Qing; Yu, Yang-Yang; Guo, Jian; Wang, Kun; Xing, Bao-Cai; Zheng, Qing-Feng; Campa, Michael J.; Patz, Edward F.; Li, Shi-You; He, You-Wen

2016-01-01

In contrast to its inhibitory effects on many cells, IL-10 activates CD8+ tumor infiltrating lymphocytes (TILs) and enhances their antitumor activity. However, CD8+ TILs do not routinely express IL-10 as autocrine complement C3 inhibits IL-10 production through complement receptors C3aR and C5aR. CD8+ TILs from C3-deficient mice, however, express IL-10 and exhibit enhanced effector function. C3-deficient mice are resistant to tumor development in a T cell- and IL-10-dependent manner; human TILs expanded with IL-2 plus IL-10 increase the killing of primary tumors in vitro compared to IL-2 treated TILs. Complement-mediated inhibition of antitumor immunity is independent of the PD-1/PD-L1 immune checkpoint pathway. Our findings suggest that complement receptors C3aR and C5aR expressed on CD8+ TILs represent a novel class of immune checkpoints that could be targeted for tumor immunotherapy. Moreover, incorporation of IL-10 in the expansion of TILs and in gene-engineered T cells for adoptive cell therapy enhances their antitumor efficacy. PMID:27297552

Bactericidal activity of glutaraldehyde-like compounds from olive products.

PubMed

Medina, Eduardo; Brenes, Manuel; García, Aranzazu; Romero, Concepción; de Castro, Antonio

2009-12-01

The bactericidal effects of several olive compounds (nonenal, oleuropein, tyrosol, the dialdehydic form of decarboxymethyl elenolic acid either free [EDA] or linked to tyrosol [TyEDA] or to hydroxytyrosol [HyEDA]), other food phenolic compounds (catechin, epicatechin, eugenol, thymol, carvacrol, and carnosic acid), and commercial disinfectants (glutaraldehyde [GTA] and ortho-phthalaldehyde [OPA]), were tested against strains of Pseudomonas fluorescens, Staphylococcus aureus, Enterococcus faecalis, and Escherichia coli. It was found that the bactericidal activities of olive GTA-like compounds (EDA, HyEDA, and TyEDA) were greater than those exerted by several food phenolic substances. Surprisingly, these olive antimicrobials were as active as the synthetic biocides GTA and OPA against the four bacteria studied. Thus, it has been proposed that the bactericidal activity of the main olive antimicrobials is primarily due to their dialdehydic structure, which is similar to that of the commercial biocides GTA and OPA. Our results clearly reveal that olive GTA-like compounds possess a strong bactericidal activity even greater than that of other food phenolic compounds or synthetic biocides.

Immunity to Escherichia coli in pigs: Serum Gamma Globulin Levels, Indirect Hemagglutinating Antibody Titres and Bactericidal Activity Against E. coli in pigs up to five Weeks of Age

PubMed Central

Wilson, M. R.; Svendsen, J.

1972-01-01

Serum gamma globulin levels, indirect hemagglutinating antibody titres and bactericidal activity against the 0149:K91;K88ac:H10 Serotype of Escherichia coli were determined in pigs up to five weeks of age from vaccinated and non-vaccinated sows. Gamma globulin levels at two days of age were approximately twice adult levels, by three weeks of age they were one quarter of adult levels and remained so until five weeks of age. Indirect hemagglutinating antibody activity was highest at two days of age, fell until three weeks of age and then rose. Little or no indirect hemagglutinating antibody activity was detected in sera taken at two days of age from pigs from non-vaccinated sows. Only three of 26 two day old pigs had demonstrable bactericidal activity; by three weeks of age 16 of 26 had bactericidal activity. Serum from piglets of vaccinated sows had no more bactericidal activity than did sera from non-vaccinated sows. PMID:4110608

An amphioxus gC1q protein binds human IgG and initiates the classical pathway: Implications for a C1q-mediated complement system in the basal chordate.

PubMed

Gao, Zhan; Li, Mengyang; Ma, Jie; Zhang, Shicui

2014-12-01

The origin of the classical complement pathway remains open during chordate evolution. A C1q-like member, BjC1q, was identified in the basal chordate amphioxus. It is predominantly expressed in the hepatic caecum, hindgut, and notochord, and is significantly upregulated following challenge with bacteria or lipoteichoic acid and LPS. Recombinant BjC1q and its globular head domain specifically interact with lipoteichoic acid and LPS, but BjC1q displays little lectin activity. Moreover, rBjC1q can assemble to form the high molecular weight oligomers necessary for binding to proteases C1r/C1s and for complement activation, and binds human C1r/C1s/mannan-binding lectin-associated serine protease-2 as well as amphioxus serine proteases involved in the cleavage of C4/C2, and C3 activation. Importantly, rBjC1q binds with human IgG as well as an amphioxus Ig domain containing protein, resulting in the activation of the classical complement pathway. This is the first report showing that a C1q-like protein in invertebrates is able to initiate classical pathway, raising the possibility that amphioxus possesses a C1q-mediated complement system. It also suggests a new scenario for the emergence of the classical complement pathway, in contrast to the proposal that the lectin pathway evolved into the classical pathway. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The dlt genes play a role in antimicrobial tolerance of Streptococcus mutans biofilms.

PubMed

Nilsson, Martin; Rybtke, Morten; Givskov, Michael; Høiby, Niels; Twetman, Svante; Tolker-Nielsen, Tim

2016-09-01

Microbial biofilms are tolerant to antibiotic treatment and therefore cause problematic infections. Knowledge about the molecular mechanisms underlying biofilm-associated antimicrobial tolerance will aid the development of antibiofilm drugs. Screening of a Streptococcus mutans transposon mutant library for genes that are important for biofilm-associated antimicrobial tolerance provided evidence that the dlt genes play a role in the tolerance of S. mutans biofilms towards gentamicin. The minimum bactericidal concentration for biofilm cells (MBC-B) for a dltA transposon mutant was eight-fold lower than that of the wild-type. The minimum bactericidal concentration for planktonic cells (MBC-P) was only slightly reduced, indicating that the mechanism involved in the observed antimicrobial tolerance has a predominant role specifically in biofilms. Experiments with a knockout dltA mutant and complemented strain confirmed that the dlt genes in S. mutans play a role in biofilm-associated tolerance to gentamicin. Confocal laser scanning microscopy analyses of biofilms grown on glass slides showed that the dltA mutant produced roughly the same amount of biofilm as the wild-type, indicating that the reduced antimicrobial tolerance of the dltA mutant is not due to a defect in biofilm formation. The products of the dlt genes have been shown to mediate alanylation of teichoic acids, and in accordance the dltA mutant showed a more negatively charged surface than the wild-type, which likely is an important factor in the reduced tolerance of the dltA mutant biofilms towards the positively charged gentamicin. Copyright © 2016 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

Discriminating the hemolytic risk of blood type A plasmas using the complement hemolysis using human erythrocytes (CHUHE) assay.

PubMed

Cunnion, Kenji M; Hair, Pamela S; Krishna, Neel K; Sass, Megan A; Enos, Clinton W; Whitley, Pamela H; Maes, Lanne Y; Goldberg, Corinne L

2017-03-01

The agglutination-based cross-matching method is sensitive for antibody binding to red blood cells but is only partially predictive of complement-mediated hemolysis, which is important in many acute hemolytic transfusion reactions. Here, we describe complement hemolysis using human erythrocytes (CHUHE) assays that directly evaluate complement-mediated hemolysis between individual serum-plasma and red blood cell combinations. The CHUHE assay is used to evaluate correlations between agglutination titers and complement-mediated hemolysis as well as the hemolytic potential of plasma from type A blood donors. Plasma or serum from each type A blood donor was incubated with AB or B red blood cells in the CHUHE assay and measured for free hemoglobin release. CHUHE assays for serum or plasma demonstrate a wide, dynamic range and high sensitivity for complement-mediated hemolysis for individual serum/plasma and red blood cell combinations. CHUHE results suggest that agglutination assays alone are only moderately predictive of complement-mediated hemolysis. CHUHE results also suggest that plasma from particular type A blood donors produce minimal complement-mediated hemolysis, whereas plasma from other type A blood donors produce moderate to high-level complement-mediated hemolysis, depending on the red blood cell donor. The current results indicate that the CHUHE assay can be used to assess complement-mediated hemolysis for plasma or serum from a type A blood donor, providing additional risk discrimination over agglutination titers alone. © 2016 AABB.

Antibiotic efficacy is linked to bacterial cellular respiration

PubMed Central

Lobritz, Michael A.; Belenky, Peter; Porter, Caroline B. M.; Gutierrez, Arnaud; Yang, Jason H.; Schwarz, Eric G.; Dwyer, Daniel J.; Khalil, Ahmad S.; Collins, James J.

2015-01-01

Bacteriostatic and bactericidal antibiotic treatments result in two fundamentally different phenotypic outcomes—the inhibition of bacterial growth or, alternatively, cell death. Most antibiotics inhibit processes that are major consumers of cellular energy output, suggesting that antibiotic treatment may have important downstream consequences on bacterial metabolism. We hypothesized that the specific metabolic effects of bacteriostatic and bactericidal antibiotics contribute to their overall efficacy. We leveraged the opposing phenotypes of bacteriostatic and bactericidal drugs in combination to investigate their activity. Growth inhibition from bacteriostatic antibiotics was associated with suppressed cellular respiration whereas cell death from most bactericidal antibiotics was associated with accelerated respiration. In combination, suppression of cellular respiration by the bacteriostatic antibiotic was the dominant effect, blocking bactericidal killing. Global metabolic profiling of bacteriostatic antibiotic treatment revealed that accumulation of metabolites involved in specific drug target activity was linked to the buildup of energy metabolites that feed the electron transport chain. Inhibition of cellular respiration by knockout of the cytochrome oxidases was sufficient to attenuate bactericidal lethality whereas acceleration of basal respiration by genetically uncoupling ATP synthesis from electron transport resulted in potentiation of the killing effect of bactericidal antibiotics. This work identifies a link between antibiotic-induced cellular respiration and bactericidal lethality and demonstrates that bactericidal activity can be arrested by attenuated respiration and potentiated by accelerated respiration. Our data collectively show that antibiotics perturb the metabolic state of bacteria and that the metabolic state of bacteria impacts antibiotic efficacy. PMID:26100898

Bactericidal Activity of Octenidine to Various Genospecies of Borrelia burgdorferi, Sensu Lato Spirochetes in Vitro and in Vivo.

PubMed

Tylewska-Wierzbanowska, Stanisława; Rogulska, Urszula; Lewandowska, Grażyna; Chmielewski, Tomasz

2017-07-06

The aim of our studies was to invent a reliable method for detection of the bactericidal activity of disinfectants against Borrelia burgdorferi in suspension (in vitro) and in cell line cultures (in vivo). In the suspension method, 0.01% octenidine at 20°C and 35°C was bactericidal to Borrelia afzeli; Borrelia garini, B. burgdorferi sensu stricto after 5 minutes treatment. Increase of the temperature to 35°C speed up the bactericidal effect to 1 minute. The bactericidal action of octenidine towards B. burgdorferi spirochetes growing in fibroblasts was less effective and needed a longer time to kill them than in the suspension.

Bactericidal Activity and Postantibiotic Effect of Levofloxacin against Anaerobes

PubMed Central

Pendland, Susan L.; Diaz-Linares, Mariela; Garey, Kevin W.; Woodward, Jennifer G.; Ryu, Seonyoung; Danziger, Larry H.

1999-01-01

The bactericidal activity and postantibiotic effect (PAE) of levofloxacin against nine anaerobes were determined. Levofloxacin at concentrations of the MIC and twice the MIC was bactericidal at 24 h to five of nine and nine of nine strains, respectively. The PAE of levofloxacin following a 2-h exposure ranged from 0.06 to 2.88 h. PMID:10508042

Functional Characterization of LcpA, a Surface-Exposed Protein of Leptospira spp. That Binds the Human Complement Regulator C4BP▿

PubMed Central

Barbosa, Angela S.; Monaris, Denize; Silva, Ludmila B.; Morais, Zenaide M.; Vasconcellos, Sílvio A.; Cianciarullo, Aurora M.; Isaac, Lourdes; Abreu, Patricia A. E.

2010-01-01

We have previously shown that pathogenic leptospiral strains are able to bind C4b binding protein (C4BP). Surface-bound C4BP retains its cofactor activity, indicating that acquisition of this complement regulator may contribute to leptospiral serum resistance. In the present study, the abilities of seven recombinant putative leptospiral outer membrane proteins to interact with C4BP were evaluated. The protein encoded by LIC11947 interacted with this human complement regulator in a dose-dependent manner. The cofactor activity of C4BP bound to immobilized recombinant LIC11947 (rLIC11947) was confirmed by detecting factor I-mediated cleavage of C4b. rLIC11947 was therefore named LcpA (for leptospiral complement regulator-acquiring protein A). LcpA was shown to be an outer membrane protein by using immunoelectron microscopy, cell surface proteolysis, and Triton X-114 fractionation. The gene coding for LcpA is conserved among pathogenic leptospiral strains. This is the first characterization of a Leptospira surface protein that binds to the human complement regulator C4BP in a manner that allows this important regulator to control complement system activation mediated either by the classical pathway or by the lectin pathway. This newly identified protein may play a role in immune evasion by Leptospira spp. and may therefore represent a target for the development of a human vaccine against leptospirosis. PMID:20404075

Susceptibility of Meningococcal Strains Responsible for Two Serogroup B Outbreaks on U.S. University Campuses to Serum Bactericidal Activity Elicited by the MenB-4C Vaccine.

PubMed

Rossi, Raffaella; Beernink, Peter T; Giuntini, Serena; Granoff, Dan M

2015-12-01

In 2013 and 2014, two U.S. universities had meningococcal serogroup B outbreaks (a total of 14 cases) caused by strains from two different clonal complexes. To control the outbreaks, students were immunized with a serogroup B meningococcal vaccine (Novartis) that was not yet licensed in the United States. The vaccine (referred to as MenB-4C) contains four components capable of eliciting bactericidal activity. Both outbreak strains had high expression levels of two of the vaccine antigens (subfamily B factor H binding protein [FHbp] and neisserial heparin binding antigen [NHba]); the university B outbreak strain also had moderate expression of a third antigen, NadA. We investigated the bactericidal activity of sera from mice immunized with FHbp, NHba, or NadA and sera from MenB-4C-immunized infant macaques and an adult human. The postimmunization bactericidal activity of the macaque or human serum against isolates from university B with FHbp identification (ID) 1 that exactly matched the vaccine FHbp sequence variant was 8- to 21-fold higher than that against isolates from university A with FHbp ID 276 (96% identity to the vaccine antigen). Based on the bactericidal activity of mouse antisera to FHbp, NadA, or NHba and macaque or human postimmunization serum that had been depleted of anti-FHbp antibody, the bactericidal activity against both outbreak strains largely or entirely resulted from antibodies to FHbp. Thus, despite the high level of strain expression of FHbp from a subfamily that matched the vaccine antigen, there can be large differences in anti-FHbp bactericidal activity induced by MenB-4C vaccination. Further, strains with moderate to high NadA and/or NHba expression can be resistant to anti-NadA or anti-NHba bactericidal activity elicited by MenB-4C vaccination. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Visceral perfusion abnormalities following complement activation. Clues to the mediators of organ ischemia in trauma and sepsis. First place winner: Conrad Jobst Award.

PubMed

Schirmer, W J; Schirmer, J M; Naff, G B; Fry, D E

1988-12-01

Complement, activated during infection and injury, has been implicated as a mediator of microvascular injury and obstruction. This study examines how two potent activators of complement, zymosan, and cobra venom factor (CVF), affect systemic and visceral perfusion. Rats were injected with either saline (1 ml/kg), zymosan (5 mg/kg) or CVF (5 units/kg) at t = 0 and 30 minutes. Thermodilution cardiac output, mean arterial pressure, heart rate, systemic vascular resistance, and hematocrit were determined at t = 2 hours. Effective hepatic and renal blood flows, by clearance of galactose and p-aminohippurate respectively, were determined over the next hour. The per cent change in total hemolytic complement from t = 0 to t = 3 hours was determined by immune hemolysis of sheep erythrocytes. There was no difference in systemic hemodynamic parameters between the three groups. Hepatic blood flow was depressed in both the zymosan (3.83 +/- 0.23 ml/min/100 g) and CVF (3.72 +/- 0.20 ml/min/100 g) groups compared with controls (4.62 +/- 0.19 ml/min/100 g, P less than 0.05). Renal blood flow in the zymosan-treated group (6.40 +/- 0.24 ml/min/100 g) increased over control (4.80 +/- 0.40 ml/min/100 g, P less than 0.05) but was unchanged in the CVF group (5.06 +/- 0.23 ml/min/100 g). The amount of complement activated correlated with the change in hepatic (r = -0.419, P less than 0.05) but not renal (r = -0.008, P = 0.917) flow. Complement activation may occupy a proximal position in the pathogenesis of hepatic ischemia associated with trauma and sepsis.

Eculizumab treatment and impaired opsonophagocytic killing of meningococci by whole blood from immunized adults.

PubMed

Konar, Monica; Granoff, Dan M

2017-08-17

Eculizumab, a humanized anti-complement C5 monoclonal antibody (mAb) for treatment of paroxysmal nocturnal hemoglobinuria (PNH) and atypical hemolytic uremic syndrome, blocks the terminal complement pathway required for serum bactericidal activity (SBA). Because treated patients are at >1000-fold increased risk of meningococcal disease, vaccination is recommended; whether vaccination can protect by opsonophagocytic activity in the absence of SBA is not known. Meningococci were added to anticoagulated blood from 12 healthy adults vaccinated with meningococcal serogroup B and serogroup A, C, W, Y vaccines. Bacterial survival was measured after 3-hour incubation in the presence of eculizumab or control complement factor D inhibitor ACH-4471, which blocks the complement alternative pathway (AP) and is in phase 2 development for treatment of PNH. In the absence of inhibitors, colony formation units (CFUs) per milliliter in blood from all 12 immunized subjects decreased from ∼4000 at time 0 to sterile cultures at 3 hours. In the presence of eculizumab, there was a >22-fold increase in geometric mean CFUs per milliliter (90 596 and 114 683 CFU/mL for serogroup B and C strains, respectively; P < .0001 compared with time 0). In the presence of ACH-4471, there was a >12-fold decrease (23 and 331 CFU/mL, respectively; P < .0001). The lack of meningococci killing by blood containing eculizumab resulted from inhibition of release of C5a, a C5 split product needed for upregulation of phagocytosis. The results provide an explanation for the large number of cases of meningococcal disease in immunized patients being treated with eculizumab and suggest that vaccination may provide better protection against meningococcal disease in patients treated with an AP-specific inhibitor. © 2017 by The American Society of Hematology.

SALO, a novel classical pathway complement inhibitor from saliva of the sand fly Lutzomyia longipalpis

PubMed Central

Ferreira, Viviana P.; Fazito Vale, Vladimir; Pangburn, Michael K.; Abdeladhim, Maha; Ferreira Mendes-Sousa, Antonio; Coutinho-Abreu, Iliano V.; Rasouli, Manoochehr; Brandt, Elizabeth A.; Meneses, Claudio; Lima, Kolyvan Ferreira; Nascimento Araújo, Ricardo; Horácio Pereira, Marcos; Kotsyfakis, Michalis; Oliveira, Fabiano; Kamhawi, Shaden; Ribeiro, Jose M. C.; Gontijo, Nelder F.; Collin, Nicolas; Valenzuela, Jesus G.

2016-01-01

Blood-feeding insects inject potent salivary components including complement inhibitors into their host’s skin to acquire a blood meal. Sand fly saliva was shown to inhibit the classical pathway of complement; however, the molecular identity of the inhibitor remains unknown. Here, we identified SALO as the classical pathway complement inhibitor. SALO, an 11 kDa protein, has no homology to proteins of any other organism apart from New World sand flies. rSALO anti-complement activity has the same chromatographic properties as the Lu. longipalpis salivary gland homogenate (SGH)counterparts and anti-rSALO antibodies blocked the classical pathway complement activity of rSALO and SGH. Both rSALO and SGH inhibited C4b deposition and cleavage of C4. rSALO, however, did not inhibit the protease activity of C1s nor the enzymatic activity of factor Xa, uPA, thrombin, kallikrein, trypsin and plasmin. Importantly, rSALO did not inhibit the alternative or the lectin pathway of complement. In conclusion our data shows that SALO is a specific classical pathway complement inhibitor present in the saliva of Lu. longipalpis. Importantly, due to its small size and specificity, SALO may offer a therapeutic alternative for complement classical pathway-mediated pathogenic effects in human diseases. PMID:26758086

Neisseria meningitidis: pathogenesis and immunity.

PubMed

Pizza, Mariagrazia; Rappuoli, Rino

2015-02-01

The recent advances in cellular microbiology, genomics, and immunology has opened new horizons in the understanding of meningococcal pathogenesis and in the definition of new prophylactic intervention. It is now clear that Neissera meningitidis has evolved a number of surface structures to mediate interaction with host cells and a number of mechanisms to subvert the immune system and escape complement-mediated killing. In this review we report the more recent findings on meningococcal adhesion and on the bacteria-complement interaction highlighting the redundancy of these mechanisms. An effective vaccine against meningococcus B, based on multiple antigens with different function, has been recently licensed. The antibodies induced by the 4CMenB vaccine could mediate bacterial killing by activating directly the classical complement pathway or, indirectly, by preventing binding of fH on the bacterial surface and interfering with colonization. Copyright © 2014 The Author. Published by Elsevier Ltd.. All rights reserved.

Effect of tilmicosin on chemotactic, phagocytic, and bactericidal activities of bovine and porcine alveolar macrophages.

PubMed

Brumbaugh, Gordon W; Herman, James D; Clancy, Julianne S; Burden, Kyland I; Barry, Tracie; Simpson, R B; López, Hector Sumano

2002-01-01

To evaluate chemotactic, phagocytic, and bactericidal activities of bovine and porcine alveolar macrophages (AM) exposed to tilmicosin. 12 healthy calves and 12 healthy pigs. Lungs were obtained immediately after euthanasia; AM were collected by means of bronchoalveolar lavage and density gradient centrifugation. Chemotactic activity was evaluated by exposing AM to lipopolysaccharide or macrophage inhibitory peptide during incubation with tilmicosin. Phagocytic activity was evaluated by incubating AM with tilmicosin for 24 hours and then with tilmicosin-resistant Salmonella serotype Typhimurium. Bactericidal activity was evaluated by incubating AM with tilmicosin (0, 10, or 20 microg/ml for bovine AM; 0 or 10 microg/ml or 10 microg/ml but washed free of tilmicosin for porcine AM) and then with Mannheimia haemolytica (bovine AM) or with Actinobacillus pleuropneumoniae or Pasteurella multocida (porcine AM). Tilmicosin had no significant effects on chemotactic or phagocytic activities of bovine or porcine AM. The time-course of bactericidal activity was best described by polynomial equations. Time to cessation of bacterial growth and area under the time versus bacterial number curve were significantly affected by incubation of AM with tilmicosin. Results show that bactericidal activity of bovine and porcine AM was enhanced by tilmicosin, but not in proportion to the reported ability of AM to concentrate tilmicosin intracellularly. With or without exposure to tilmicosin, the time-course of bactericidal activity of bovine AM against M haemolytica and of porcine AM against A pleuropneumoniae or P multocida was too complex to be reduced to a simple linear equation.

Novel Evasion Mechanisms of the Classical Complement Pathway

PubMed Central

Garcia, Brandon L.; Zwarthoff, Seline A.; Rooijakkers, Suzan H. M.; Geisbrecht, Brian V.

2016-01-01

Complement is a network of soluble and cell surface-associated proteins which gives rise to a self-amplifying, yet tightly regulated system with fundamental roles in immune surveillance and clearance. Complement becomes activated on the surface of ‘non-self’ cells by one of three initiating mechanisms known as the classical, lectin, or alternative pathways. Evasion of complement function is a hallmark of invasive pathogens and hematophagous organisms. While many complement inhibition strategies hinge on hijacking activities of endogenous complement regulatory proteins, an increasing number of uniquely evolved evasion molecules have been discovered over the past decade. In this review we focus on several recent investigations which have revealed mechanistically distinct inhibitors of the classical pathway. Because the classical pathway is an important and specific mediator of various autoimmune and inflammatory disorders, in-depth knowledge of novel evasion mechanisms could direct future development of therapeutic anti-inflammatory molecules. PMID:27591336

Novel Evasion Mechanisms of the Classical Complement Pathway.

PubMed

Garcia, Brandon L; Zwarthoff, Seline A; Rooijakkers, Suzan H M; Geisbrecht, Brian V

2016-09-15

Complement is a network of soluble and cell surface-associated proteins that gives rise to a self-amplifying, yet tightly regulated system with fundamental roles in immune surveillance and clearance. Complement becomes activated on the surface of nonself cells by one of three initiating mechanisms known as the classical, lectin, and alternative pathways. Evasion of complement function is a hallmark of invasive pathogens and hematophagous organisms. Although many complement-inhibition strategies hinge on hijacking activities of endogenous complement regulatory proteins, an increasing number of uniquely evolved evasion molecules have been discovered over the past decade. In this review, we focus on several recent investigations that revealed mechanistically distinct inhibitors of the classical pathway. Because the classical pathway is an important and specific mediator of various autoimmune and inflammatory disorders, in-depth knowledge of novel evasion mechanisms could direct future development of therapeutic anti-inflammatory molecules. Copyright © 2016 by The American Association of Immunologists, Inc.

In vitro antibody-enzyme conjugates with specific bactericidal activity.

PubMed

Knowles, D M; Sulivan, T J; Parker, C W; Williams, R C

1973-06-01

IgG with antibacterial antibody opsonic activity was isolated from rabbit antisera produced by intravenous hyperimmunization with several test strains of pneumococci, Group A beta-hemolytic streptococci, Staphylococcus aureus, Proteus mirabilis, Pseudomonas aeruginosa, and Escherichia coli. Antibody-enzyme conjugates were prepared, using diethylmalonimidate to couple glucose oxidase to IgG antibacterial antibody preparations. Opsonic human IgG obtained from serum of patients with subacute bacterial endocarditis was also conjugated to glucose oxidase. Antibody-enzyme conjugates retained combining specificity for test bacteria as demonstrated by indirect immunofluorescence. In vitro test for bactericidal activity of antibody-enzyme conjugates utilized potassium iodide, lactoperoxidase, and glucose as cofactors. Under these conditions glucose oxidase conjugated to antibody generates hydrogen peroxide, and lactoperoxidase enzyme catalyzes the reduction of hydrogen peroxide with simultaneous oxidation of I(-) and halogenation and killing of test bacteria. Potent in vitro bactericidal activity of this system was repeatedly demonstrated for antibody-enzyme conjugates against pneumococci, streptococci, S. aureus, P. mirabilis, and E. coli. However, no bactericidal effect was demonstrable with antibody-enzyme conjugates and two test strains of P. aeruginosa. Bactericidal activity of antibody-enzyme conjugates appeared to parallel original opsonic potency of unconjugated IgG preparations. Antibody-enzyme conjugates at concentrations as low as 0.01 mg/ml were capable of intense bactericidal activity producing substantial drops in surviving bacterial counts within 30-60 min after initiation of assay. These in vitro bactericidal systems indicate that the concept of antibacterial antibody-enzyme conjugates may possibly be adaptable as a mechanism for treatment of patients with leukocyte dysfunction or fulminant bacteremia.

Platelet-activating factor mediates monocyte chemoattractant protein-1 expression in glomerular immune injury.

PubMed

Jocks, T; Freudenberg, J; Zahner, G; Stahl, R A

1998-01-01

These studies were designed to determine the possible role of platelet-activating factor (PAF) in the production of monocyte chemoattractant protein-1 (MCP-1) in glomerular immune injury. The glomerular lesion was induced in isolated perfused rat kidneys by a rabbit anti-rat-thymocyte serum (ATS) and rat serum (RS) as a complement source. Perfusion of kidneys with ATS and RS results in the selective binding of the antiserum to the glomerular mesangium with consecutive intraglomerular activation of complement. Antibody binding and complement activation induced a significant increase in glomerular MCP-1 mRNA levels when assessed by Northern blotting or RT-PCR. Decomplemented RS or non antibody rabbit IgG had only moderate effects on glomerular MCP-1 mRNA levels. The PAF receptor antagonist WEB 2170 almost completely blocked the ATS and RS induced MCP-1 mRNA levels. Perfusion of control kidneys with PAF increased MCP-1 mRNA expression, an effect which was blocked by WEB 2170. Glomerular MCP-1 protein formation, assessed by Western blotting, was stimulated following ATS and RS and PAF, respectively, was blocked by WEB 2170. These data show that PAF, derived from glomerular resident cells following antibody and complement induced injury, stimulates MCP-1 expression. In addition to the direct effects on leukocyte adhesion and activation PAF may mediate inflammatory cell influx in glomerular injuries due to the release of MCP-1.

Purification and characterization of a bacteriocin from an oenological strain of Leuconostoc mesenteroides subsp. cremoris.

PubMed

Dündar, Halil; Salih, Bekir; Bozoğlu, Faruk

2016-05-18

Malolactic fermentation (MLF), which improves organoleptic properties and biologic stability of some wines, may cause wine spoilage if uncontrolled. Bacteriocins were reported as efficient preservatives to control MLF through their bactericidal effect on malolactic bacteria. Leuconostoc mesenteroides subsp. cremoris W3 isolated from wine produces an inhibitory substance that is bactericidal against malolactic bacteria in model wine medium. Treatment of the culture supernatant of strain W3 with proteases eliminated the inhibitory activity, which proved that it is a true bacteriocin and we tentatively termed it mesentericin W3. The bacteriocin inhibited the growth of food-borne pathogenic bacteria such as Enterococcus faecalis, Listeria monocytogenes, and malolactic bacteria. It was active over a wide pH range and stable to organic solvents and heat. Mesentericin W3 was purified to homogeneity by a pH-mediated cell adsorption-desorption method, cation exchange, hydrophobic interaction, and reverse-phase chromatography. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectroscopy (MS) and partial amino acid sequence analysis revealed that mesentericin W3 was identical to mesentericin Y105.

Complement activation and liver impairment in trichloroethylene-sensitized BALB/c mice.

PubMed

Zhang, Jiaxiang; Zha, Wansheng; Wang, Feng; Jiang, Tao; Xu, Shuhai; Yu, Junfeng; Zhou, Chengfan; Shen, Tong; Wu, Changhao; Zhu, Qixing

2013-01-01

Our recent studies have shown that trichloroethylene (TCE) was able to induce multisystem injuries in the form of occupational medicamentosa-like dermatitis, including skin, kidney, and liver damages. However, the role of complement activation in the immune-mediated liver injury is not known. This study examined the role of complement activation in the liver injury in a mouse model of TCE-induced sensitization. Treatment of female BALB/c mice with TCE under specific dosing protocols resulted in skin inflammation and sensitization. Skin edema and erythema occurred in TCE-sensitized groups. Trichloroethylene sensitization produced liver histopathological lesions, increased serum alanine aminotransferase, aspartate transaminase activities, and the relative liver weight. The concentrations of serum complement components C3a-desArg, C5a-desArg, and C5b-9 were significantly increased in 24-hour, 48-hour, and 72-hour sensitization-positive groups treated with TCE and peaked in the 72-hour sensitization-positive group. Depositions of C3a, C5a, and C5b-9 into the liver tissue were also revealed by immunohistochemistry. Immunofluorescence further verified high C5b-9 expression in 24-hour, 48-hour, and 72-hour sensitization-positive groups in response to TCE treatment. Reverse transcription-polymerase chain reaction detected C3 messenger RNA expression in the liver, and this was significantly increased in 24-hour and 48-hour sensitization-positive groups with a transient reduction at 72 hours. These results provide the first experimental evidence that complement activation may play a key role in the generation and progression of immune-mediated hepatic injury by exposure to TCE.

Complement evasion by Bordetella pertussis: implications for improving current vaccines.

PubMed

Jongerius, Ilse; Schuijt, Tim J; Mooi, Frits R; Pinelli, Elena

2015-04-01

Bordetella pertussis causes whooping cough or pertussis, a highly contagious disease of the respiratory tract. Despite high vaccination coverage, reported cases of pertussis are rising worldwide and it has become clear that the current vaccines must be improved. In addition to the well-known protective role of antibodies and T cells during B. pertussis infection, innate immune responses such as the complement system play an essential role in B. pertussis killing. In order to evade this complement activation and colonize the human host, B. pertussis expresses several molecules that inhibit complement activation. Interestingly, one of the known complement evasion proteins, autotransporter Vag8, is highly expressed in the recently emerged B. pertussis isolates. Here, we describe the current knowledge on how B. pertussis evades complement-mediated killing. In addition, we compare this to complement evasion strategies used by other bacterial species. Finally, we discuss the consequences of complement evasion by B. pertussis on adaptive immunity and how identification of the bacterial molecules and the mechanisms involved in complement evasion might help improve pertussis vaccines.

Understanding bactericidal performance on ambient light activated TiO2-InVO4 nanostructured films.

PubMed

He, Ziming; Xu, Qingchi; Tan, Timothy Thatt Yang

2011-12-01

TiO(2)-InVO(4) nanostructured films were coated onto glass substrates and systematically investigated for their bactericidal activities using Escherichia coli (E. coli) as the model bacterium under ambient light illumination. The uniform TiO(2)-InVO(4) nanostructured films were prepared using titanium isopropoxide (TTIP) as the precursor via a simple sol-gel approach. Polyethylenimine (PEI) was used as a surfactant to ensure uniform dispersion of InVO(4) and a sacrificial pore-inducing agent, generating nanostructured films. Compared to unmodified TiO(2) film, the current TiO(2)-InVO(4) films exhibited enhanced bactericidal activities under ambient light illumination. Bacterial cell "photo-fixation" was demonstrated to be crucial in enhancing the bactericidal activity. A bacterial-nanostructured surface interaction mechanism was proposed for the current ambient-light activated nanostructured film.

Small-molecule factor D inhibitors selectively block the alternative pathway of complement in paroxysmal nocturnal hemoglobinuria and atypical hemolytic uremic syndrome.

PubMed

Yuan, Xuan; Gavriilaki, Eleni; Thanassi, Jane A; Yang, Guangwei; Baines, Andrea C; Podos, Steven D; Huang, Yongqing; Huang, Mingjun; Brodsky, Robert A

2017-03-01

Paroxysmal nocturnal hemoglobinuria and atypical hemolytic uremic syndrome are diseases of excess activation of the alternative pathway of complement that are treated with eculizumab, a humanized monoclonal antibody against the terminal complement component C5. Eculizumab must be administered intravenously, and moreover some patients with paroxysmal nocturnal hemoglobinuria on eculizumab have symptomatic extravascular hemolysis, indicating an unmet need for additional therapeutic approaches. We report the activity of two novel small-molecule inhibitors of the alternative pathway component Factor D using in vitro correlates of both paroxysmal nocturnal hemoglobinuria and atypical hemolytic uremic syndrome. Both compounds bind human Factor D with high affinity and effectively inhibit its proteolytic activity against purified Factor B in complex with C3b. When tested using the traditional Ham test with cells from paroxysmal nocturnal hemoglobinuria patients, the Factor D inhibitors significantly reduced complement-mediated hemolysis at concentrations as low as 0.01 μM. Additionally the compound ACH-4471 significantly decreased C3 fragment deposition on paroxysmal nocturnal hemoglobinuria erythrocytes, indicating a reduced potential relative to eculizumab for extravascular hemolysis. Using the recently described modified Ham test with serum from patients with atypical hemolytic uremic syndrome, the compounds reduced the alternative pathway-mediated killing of PIGA -null reagent cells, thus establishing their potential utility for this disease of alternative pathway of complement dysregulation and validating the modified Ham test as a system for pre-clinical drug development for atypical hemolytic uremic syndrome. Finally, ACH-4471 blocked alternative pathway activity when administered orally to cynomolgus monkeys. In conclusion, the small-molecule Factor D inhibitors show potential as oral therapeutics for human diseases driven by the alternative pathway of complement, including paroxysmal nocturnal hemoglobinuria and atypical hemolytic uremic syndrome. Copyright© Ferrata Storti Foundation.

Hemorrhage-induced intestinal damage is complement independent in Helicobacter-hepaticus infected mice

PubMed Central

Hylton, Diana J.; Phillips, Lauren M.; Hoffman, Sara M.; Fleming, Sherry D.

2010-01-01

With over half of the world population infected, Helicobacter infection is an important public health issue associated with gastrointestinal cancers and inflammatory bowel disease. Animal studies indicate that complement and oxidative stress play a role in Helicobacter infections. Hemorrhage induces tissue damage which is attenuated by blockade of either complement activation or oxidative stress products. Therefore, we hypothesized that chronic Helicobacter hepaticus infection would modulate hemorrhage-induced intestinal damage and inflammation. To test this hypothesis, we examined hemorrhage-induced jejunal damage and inflammation in uninfected and H. hepaticus infected mice. H. hepaticus infection increased hemorrhage-induced mid-jejunal mucosal damage despite attenuating complement activation. In addition, infection alone increased chemokine secretion, changing the hemorrhage-induced neutrophil infiltration to a macrophage-mediated inflammatory response. The hemorrhage-induced macrophage infiltration correlated with increased secretion of tumor necrosis factor-α (TNF-α3) and nitric oxide (NO) in the infected mice. Together these data indicate that Helicobacter infection modulates the mechanism of hemorrhage-induced intestinal damage and inflammation from a complement-mediated response to a macrophage response with elevated TNF-α and NO. These data indicate that chronic, low level infections change the response to trauma and should be considered when designing and administering therapeutics. PMID:20220569

Soya bean Gα proteins with distinct biochemical properties exhibit differential ability to complement Saccharomyces cerevisiae gpa1 mutant.

PubMed

Roy Choudhury, Swarup; Wang, Yuqi; Pandey, Sona

2014-07-01

Signalling pathways mediated by heterotrimeric G-proteins are common to all eukaryotes. Plants have a limited number of each of the G-protein subunits, with the most elaborate G-protein network discovered so far in soya bean (Glycine max, also known as soybean) which has four Gα, four Gβ and ten Gγ proteins. Biochemical characterization of Gα proteins from plants suggests significant variation in their properties compared with the well-characterized non-plant proteins. Furthermore, the four soya bean Gα (GmGα) proteins exhibit distinct biochemical activities among themselves, but the extent to which such biochemical differences contribute to their in vivo function is also not known. We used the yeast gpa1 mutant which displays constitutive signalling and growth arrest in the pheromone-response pathway as an in vivo model to evaluate the effect of distinct biochemical activities of GmGα proteins. We showed that specific GmGα proteins can be activated during pheromone-dependent receptor-mediated signalling in yeast and they display different strengths towards complementation of yeast gpa1 phenotypes. We also identified amino acids that are responsible for differential complementation abilities of specific Gα proteins. These data establish that specific plant Gα proteins are functional in the receptor-mediated pheromone-response pathway in yeast and that the subtle biochemical differences in their activity are physiologically relevant.

Ectromelia virus inhibitor of complement enzymes protects intracellular mature virus and infected cells from mouse complement.

PubMed

Moulton, Elizabeth A; Bertram, Paula; Chen, Nanhai; Buller, R Mark L; Atkinson, John P

2010-09-01

Poxviruses produce complement regulatory proteins to subvert the host's immune response. Similar to the human pathogen variola virus, ectromelia virus has a limited host range and provides a mouse model where the virus and the host's immune response have coevolved. We previously demonstrated that multiple components (C3, C4, and factor B) of the classical and alternative pathways are required to survive ectromelia virus infection. Complement's role in the innate and adaptive immune responses likely drove the evolution of a virus-encoded virulence factor that regulates complement activation. In this study, we characterized the ectromelia virus inhibitor of complement enzymes (EMICE). Recombinant EMICE regulated complement activation on the surface of CHO cells, and it protected complement-sensitive intracellular mature virions (IMV) from neutralization in vitro. It accomplished this by serving as a cofactor for the inactivation of C3b and C4b and by dissociating the catalytic domain of the classical pathway C3 convertase. Infected murine cells initiated synthesis of EMICE within 4 to 6 h postinoculation. The levels were sufficient in the supernatant to protect the IMV, upon release, from complement-mediated neutralization. EMICE on the surface of infected murine cells also reduced complement activation by the alternative pathway. In contrast, classical pathway activation by high-titer antibody overwhelmed EMICE's regulatory capacity. These results suggest that EMICE's role is early during infection when it counteracts the innate immune response. In summary, ectromelia virus produced EMICE within a few hours of an infection, and EMICE in turn decreased complement activation on IMV and infected cells.

Cyclosporine Induces Endothelial Cell Release of Complement-Activating Microparticles

PubMed Central

Renner, Brandon; Klawitter, Jelena; Goldberg, Ryan; McCullough, James W.; Ferreira, Viviana P.; Cooper, James E.; Christians, Uwe

2013-01-01

Defective control of the alternative pathway of complement is an important risk factor for several renal diseases, including atypical hemolytic uremic syndrome. Infections, drugs, pregnancy, and hemodynamic insults can trigger episodes of atypical hemolytic uremic syndrome in susceptible patients. Although the mechanisms linking these clinical events with disease flares are unknown, recent work has revealed that each of these clinical conditions causes cells to release microparticles. We hypothesized that microparticles released from injured endothelial cells promote intrarenal complement activation. Calcineurin inhibitors cause vascular and renal injury and can trigger hemolytic uremic syndrome. Here, we show that endothelial cells exposed to cyclosporine in vitro and in vivo release microparticles that activate the alternative pathway of complement. Cyclosporine-induced microparticles caused injury to bystander endothelial cells and are associated with complement-mediated injury of the kidneys and vasculature in cyclosporine-treated mice. Cyclosporine-induced microparticles did not bind factor H, an alternative pathway regulatory protein present in plasma, explaining their complement-activating phenotype. Finally, we found that in renal transplant patients, the number of endothelial microparticles in plasma increases 2 weeks after starting tacrolimus, and treatment with tacrolimus associated with increased C3 deposition on endothelial microparticles in the plasma of some patients. These results suggest that injury-associated release of endothelial microparticles is an important mechanism by which systemic insults trigger intravascular complement activation and complement-dependent renal diseases. PMID:24092930

The Scl1 protein of M6-type group A Streptococcus binds the human complement regulatory protein, factor H, and inhibits the alternative pathway of complement.

PubMed

Caswell, Clayton C; Han, Runlin; Hovis, Kelley M; Ciborowski, Pawel; Keene, Douglas R; Marconi, Richard T; Lukomski, Slawomir

2008-02-01

Non-specific activation of the complement system is regulated by the plasma glycoprotein factor H (FH). Bacteria can avoid complement-mediated opsonization and phagocytosis through acquiring FH to the cell surface. Here, we characterize an interaction between the streptococcal collagen-like protein Scl1.6 of M6-type group A Streptococcus (GAS) and FH. Using affinity chromatography with immobilized recombinant Scl1.6 protein, we co-eluted human plasma proteins with molecular weight of 155 kDa, 43 kDa and 38 kDa. Mass spectrometry identified the 155 kDa band as FH and two other bands as isoforms of the FH-related protein-1. The identities of all three bands were confirmed by Western immunoblotting with specific antibodies. Structure-function relation studies determined that the globular domain of the Scl1.6 variant specifically binds FH while fused to collagenous tails of various lengths. This binding is not restricted to Scl1.6 as the phylogenetically linked Scl1.55 variant also binds FH. Functional analyses demonstrated the cofactor activity of the rScl1.6-bound FH for factor I-mediated cleavage of C3b. Finally, purified FH bound to the Scl1.6 protein present in the cell wall material obtained from M6-type GAS. In conclusion, we have identified a functional interaction between Scl1 and plasma FH, which may contribute to GAS evasion of complement-mediated opsonization and phagocytosis.

Evaluation of bactericidal efficacy of silver ions on Escherichia coli for drinking water disinfection.

PubMed

Pathak, Satya P; Gopal, K

2012-07-01

The purpose of this study is the development of a suitable process for the disinfection of drinking water by evaluating bactericidal efficacy of silver ions from silver electrodes. A prototype of a silver ioniser with silver electrodes and control unit has been fabricated. Silver ions from silver electrodes in water samples were estimated with an atomic absorption spectrophotometer. A fresh culture of Escherichia coli (1.75 × 10(3) c.f.u./ml) was exposed to 1, 2, 5, 10 and 20 ppb of silver ions in 100 ml of autoclaved tap water for 60 min. The effect of different pH and temperatures on bactericidal efficacy was observed at constant silver ion concentration (5 ppb) and contact time of 30 min. The maximum bactericidal activity (100%) was observed at 20 ppb of silver ion concentration indicating total disinfection after 20 min while minimum bactericidal activity (25%) was observed after 10 min at 01 ppb of silver ions. Likewise, 100% bactericidal activity was noticed with 2, 5 and 10 ppb of silver ions after 60, 50 and 40 min, respectively. Bactericidal activity at pH 5, 6, 7, 8 and 9 was observed at 79.9%, 79.8%, 80.5%, 100% and 100%, respectively, whereas it was 80.4%, 88.3%, 100%, 100% and 100% at 10°C, 20°C, 30°C, 40°C and 50°C, respectively. The findings of this study revealed that very low concentrations of silver ions at pH 8-9 and temperature >20°C have bactericidal efficacy for total disinfection of drinking water. Silver ionisation is suitable for water disinfection and an appropriate alternative to chlorination which forms carcinogenic disinfection by-products.

Effects of bacteriocins on methicillin-resistant Staphylococcus aureus biofilm.

PubMed

Okuda, Ken-ichi; Zendo, Takeshi; Sugimoto, Shinya; Iwase, Tadayuki; Tajima, Akiko; Yamada, Satomi; Sonomoto, Kenji; Mizunoe, Yoshimitsu

2013-11-01

Control of biofilms formed by microbial pathogens is an important subject for medical researchers, since the development of biofilms on foreign-body surfaces often causes biofilm-associated infections in patients with indwelling medical devices. The present study examined the effects of different kinds of bacteriocins, which are ribosomally synthesized antimicrobial peptides produced by certain bacteria, on biofilms formed by a clinical isolate of methicillin-resistant Staphylococcus aureus (MRSA). The activities and modes of action of three bacteriocins with different structures (nisin A, lacticin Q, and nukacin ISK-1) were evaluated. Vancomycin, a glycopeptide antibiotic used in the treatment of MRSA infections, showed bactericidal activity against planktonic cells but not against biofilm cells. Among the tested bacteriocins, nisin A showed the highest bactericidal activity against both planktonic cells and biofilm cells. Lacticin Q also showed bactericidal activity against both planktonic cells and biofilm cells, but its activity against biofilm cells was significantly lower than that of nisin A. Nukacin ISK-1 showed bacteriostatic activity against planktonic cells and did not show bactericidal activity against biofilm cells. Mode-of-action studies indicated that pore formation leading to ATP efflux is important for the bactericidal activity against biofilm cells. Our results suggest that bacteriocins that form stable pores on biofilm cells are highly potent for the treatment of MRSA biofilm infections.

Effects of Bacteriocins on Methicillin-Resistant Staphylococcus aureus Biofilm

PubMed Central

Zendo, Takeshi; Sugimoto, Shinya; Iwase, Tadayuki; Tajima, Akiko; Yamada, Satomi; Sonomoto, Kenji

2013-01-01

Control of biofilms formed by microbial pathogens is an important subject for medical researchers, since the development of biofilms on foreign-body surfaces often causes biofilm-associated infections in patients with indwelling medical devices. The present study examined the effects of different kinds of bacteriocins, which are ribosomally synthesized antimicrobial peptides produced by certain bacteria, on biofilms formed by a clinical isolate of methicillin-resistant Staphylococcus aureus (MRSA). The activities and modes of action of three bacteriocins with different structures (nisin A, lacticin Q, and nukacin ISK-1) were evaluated. Vancomycin, a glycopeptide antibiotic used in the treatment of MRSA infections, showed bactericidal activity against planktonic cells but not against biofilm cells. Among the tested bacteriocins, nisin A showed the highest bactericidal activity against both planktonic cells and biofilm cells. Lacticin Q also showed bactericidal activity against both planktonic cells and biofilm cells, but its activity against biofilm cells was significantly lower than that of nisin A. Nukacin ISK-1 showed bacteriostatic activity against planktonic cells and did not show bactericidal activity against biofilm cells. Mode-of-action studies indicated that pore formation leading to ATP efflux is important for the bactericidal activity against biofilm cells. Our results suggest that bacteriocins that form stable pores on biofilm cells are highly potent for the treatment of MRSA biofilm infections. PMID:23979748

Persistence of antibodies in adolescents 18−24 months after immunization with one, two, or three doses of 4CMenB meningococcal serogroup B vaccine

PubMed Central

Santolaya, Maria Elena; O'Ryan, Miguel; Valenzuela, María Teresa; Prado, Valeria; Vergara, Rodrigo F; Muñoz, Alma; Toneatto, Daniela; Graña, Gabriela; Wang, Huajun; Dull, Peter M

2013-01-01

We previously demonstrated the immunogenicity and tolerability of the serogroup B meningococcal vaccine, 4CMenB (Bexsero®), in 11−17 y-olds randomized to receive 1, 2, or 3 doses at 1, 2, or 6 mo intervals. Participants in this extension study provided an additional blood sample 18−24 mo after last vaccine dose, to assess persistence of serum bactericidal activity with human complement (hSBA), and to compare with age-matched 4CMenB-naïve controls. In the original study, one month after one 4CMenB dose, 93% of subjects had seroprotective hSBA titers (≥4) against indicator serogroup B strains for individual vaccine antigens (fHbp, NadA and NZOMV), increasing to ~100% after two or three doses. After 18−24 mo, 62−73% of subjects given one dose had titers ≥4 against the three antigens, significantly lower rates than after two (77−94%) or three (86−97%) doses. Only proportions with titers ≥ 4 against NZOMV were significantly different between the two (77%) and three (90%, p < 0.0001) dose groups. These results confirm that two doses of 4CMenB, administered 1 to 6 mo apart, provide good levels of bactericidal activity against serogroup B meningococci, which were sustained at least 18−24 mo in over 64% of adolescents for all three tested vaccine-related antigens. PMID:23811804

SIGN-R1 and complement factors are involved in the systemic clearance of radiation-induced apoptotic cells in whole-body irradiated mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Jin-Yeon; Loh, SoHee; Cho, Eun-hee

Although SIGN-R1-mediated complement activation pathway has been shown to enhance the systemic clearance of apoptotic cells, the role of SIGN-R1 in the clearance of radiation-induced apoptotic cells has not been characterized and was investigated in this study. Our data indicated that whole-body γ-irradiation of mice increased caspase-3{sup +} apoptotic lymphocyte numbers in secondary lymphoid organs. Following γ-irradiation, SIGN-R1 and complements (C4 and C3) were simultaneously increased only in the mice spleen tissue among the assessed tissues. In particular, C3 was exclusively activated in the spleen. The delayed clearance of apoptotic cells was markedly prevalent in the spleen and liver ofmore » SIGN-R1 KO mice, followed by a significant increase of CD11b{sup +} cells. These results indicate that SIGN-R1 and complement factors play an important role in the systemic clearance of radiation-induced apoptotic innate immune cells to maintain tissue homeostasis after γ-irradiation. - Highlights: • Splenic SIGN-R1{sup +} macrophages are activated after γ-irradiation. • C3 and C4 levels increased and C3 was activated in the spleen after γ-irradiation. • SIGN-R1 mediated the systemic clearance of radiation-induced apoptotic cells in spleen and liver.« less

Complement factor H protects mice from ischemic acute kidney injury but is not critical for controlling complement activation by glomerular IgM.

PubMed

Goetz, Lindsey; Laskowski, Jennifer; Renner, Brandon; Pickering, Matthew C; Kulik, Liudmila; Klawitter, Jelena; Stites, Erik; Christians, Uwe; van der Vlag, Johan; Ravichandran, Kameswaran; Holers, V Michael; Thurman, Joshua M

2018-05-01

Natural IgM binds to glomerular epitopes in several progressive kidney diseases. Previous work has shown that IgM also binds within the glomerulus after ischemia/reperfusion (I/R) but does not fully activate the complement system. Factor H is a circulating complement regulatory protein, and congenital or acquired deficiency of factor H is a strong risk factor for several types of kidney disease. We hypothesized that factor H controls complement activation by IgM in the kidney after I/R, and that heterozygous factor H deficiency would permit IgM-mediated complement activation and injury at this location. We found that mice with targeted heterozygous deletion of the gene for factor H developed more severe kidney injury after I/R than wild-type controls, as expected, but that complement activation within the glomeruli remained well controlled. Furthermore, mice that are unable to generate soluble IgM were not protected from renal I/R, even in the setting of heterozygous factor H deficiency. These results demonstrate that factor H is important for limiting injury in the kidney after I/R, but it is not critical for controlling complement activation by immunoglobulin within the glomerulus in this setting. IgM binds to glomerular epitopes after I/R, but it is not a significant source of injury. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

KINETICS OF THE ACTION OF AMPICILLIN ON ESCHERICHIA COLI

PubMed Central

Seligman, Stephen J.; Hewitt, William L.

1963-01-01

Seligman, Stephen J. (University of California, Los Angeles) and William L. Hewitt. Kinetics of the action of ampicillin on Escherichia coli. J. Bacteriol. 85:1160–1164. 1963.—The curve of the number of viable Escherichia coli after exposure to ampicillin can be divided into three phases: a lag phase, a rapid bactericidal phase, and a slow bactericidal phase. Some of the variables affecting the magnitude of the first two of these phases were investigated. Progressive lowering of drug concentration resulted in prolongation of the lag phase and decrease in slope and extent of the rapid bactericidal phase. The production of elongated gram-negative forms and the emergence of a mutant with increased penicillinase activity complicated interpretation of the lower dose curves. With sufficient drug concentration, the length of the lag phase and the slope of the rapid bactericidal curve were independent of the size of inoculum up to 108 organisms. Varying pH revealed that maximal activity, as measured by the shortest lag phase and steepest slope of the rapid bactericidal phase, was present at slightly acid pH levels. Increasing pH resulted principally in prolongation of lag phase. With greater acidity, decrease in slope of the rapid bactericidal phase was more prominent. Cultures studied under conditions of lessened metabolic activity exhibited prolonged lag phase and decreased slope and extent of rapid bactericidal phase. PMID:14044010

Enhanced protective antibody to a mutant meningococcal factor H-binding protein with low-factor H binding

PubMed Central

Granoff, Dan M.; Giuntini, Serena; Gowans, Flor A.; Lujan, Eduardo; Sharkey, Kelsey; Beernink, Peter T.

2016-01-01

Meningococcal factor H-binding protein (FHbp) is an antigen in 2 serogroup B meningococcal vaccines. FHbp specifically binds human and some nonhuman primate complement FH. To investigate the effect of binding of FH to FHbp on protective antibody responses, we immunized infant rhesus macaques with either a control recombinant FHbp antigen that bound macaque FH or a mutant antigen with 2 amino acid substitutions and >250-fold lower affinity for FH. The mutant antigen elicited 3-fold higher serum IgG anti-FHbp titers and up to 15-fold higher serum bactericidal titers than the control FHbp vaccine. When comparing sera with similar IgG anti-FHbp titers, the antibodies elicited by the mutant antigen gave greater deposition of complement component C4b on live meningococci (classical complement pathway) and inhibited binding of FH, while the anti-FHbp antibodies elicited by the control vaccine enhanced FH binding. Thus, the mutant FHbp vaccine elicited an anti-FHbp antibody repertoire directed at FHbp epitopes within the FH binding site, which resulted in greater protective activity than the antibodies elicited by the control vaccine, which targeted FHbp epitopes outside of the FH combining site. Binding of a host protein to a vaccine antigen impairs protective antibody responses, which can be overcome with low-binding mutant antigens. PMID:27668287

A randomized, first-in-human, healthy volunteer trial of BIVV009, a humanized antibody for the specific inhibition of the classical complement pathway.

PubMed

Bartko, Johann; Schoergenhofer, Christian; Schwameis, Michael; Firbas, Christa; Beliveau, Martin; Chang, Colin; Marier, Jean-Francois; Nix, Darrell; Gilbert, James C; Panicker, Sandip; Jilma, Bernd

2018-05-08

Aberrant activation of the classical complement pathway is the common underlying pathophysiology of orphan diseases such as bullous pemphigoid, antibody-mediated rejection of organ transplants, cold agglutinin disease and warm autoimmune haemolytic anaemia. Therapeutic options for these complement-mediated disorders are limited and BIVV009, a humanized monoclonal antibody directed against complement factor C1s, may be potentially useful for inhibition of the classical complement pathway. A phase-1, first-in-human, double-blind, randomized, placebo-controlled, dose-escalation trial of single and multiple doses of BIVV009 or placebo was conducted in 64 volunteers to evaluate safety, tolerability, pharmacokinetic, and pharmacodynamic profiles. Single and multiple infusions of BIVV009 were well tolerated without any safety concerns. BIVV009 exhibited a steep concentration-effect relationship with a Hill coefficient of 2.4, and an IC90 of 15.5 µg/mL. This study establishes the foundation for using BIVV009 as a highly selective inhibitor of the classical complement pathway in different diseases. This article is protected by copyright. All rights reserved. © 2018 American Society for Clinical Pharmacology and Therapeutics.

Enhanced CDC of B cell chronic lymphocytic leukemia cells mediated by rituximab combined with a novel anti-complement factor H antibody.

PubMed

Winkler, Mark T; Bushey, Ryan T; Gottlin, Elizabeth B; Campa, Michael J; Guadalupe, Eross S; Volkheimer, Alicia D; Weinberg, J Brice; Patz, Edward F

2017-01-01

Rituximab therapy for B cell chronic lymphocytic leukemia (B-CLL) has met with mixed success. Among several factors to which resistance can be attributed is failure to activate complement dependent cytotoxicity (CDC) due to protective complement regulatory proteins, including the soluble regulator complement factor H (CFH). We hypothesized that rituximab killing of non-responsive B-CLL cells could be augmented by a novel human monoclonal antibody against CFH. The B cells from 11 patients with B-CLL were tested ex vivo in CDC assays with combinations of CFH monoclonal antibody, rituximab, and a negative control antibody. CDC of rituximab non-responsive malignant B cells from CLL patients could in some cases be augmented by the CFH monoclonal antibody. Antibody-mediated cytotoxicity of cells was dependent upon functional complement. In one case where B-CLL cells were refractory to CDC by the combination of rituximab plus CFH monoclonal antibody, additionally neutralizing the membrane complement regulatory protein CD59 allowed CDC to occur. Inhibiting CDC regulatory proteins such as CFH holds promise for overcoming resistance to rituximab therapy in B-CLL.

Targeting the complement system for the management of retinal inflammatory and degenerative diseases.

PubMed

Xu, Heping; Chen, Mei

2016-09-15

The retina, an immune privileged tissue, has specialized immune defense mechanisms against noxious insults that may exist in diseases such as age-related macular degeneration (AMD), diabetic retinopathy (DR), uveoretinitis and glaucoma. The defense system consists of retinal innate immune cells (including microglia, perivascular macrophages, and a small population of dendritic cells) and the complement system. Under normal aging conditions, retinal innate immune cells and the complement system undergo a low-grade activation (parainflammation) which is important for retinal homeostasis. In disease states such as AMD and DR, the parainflammatory response is dysregulated and develops into detrimental chronic inflammation. Complement activation in the retina is an important part of chronic inflammation and may contribute to retinal pathology in these disease states. Here, we review the evidence that supports the role of uncontrolled or dysregulated complement activation in various retinal degenerative and angiogenic conditions. We also discuss current strategies that are used to develop complement-based therapies for retinal diseases such as AMD. The potential benefits of complement inhibition in DR, uveoretinitis and glaucoma are also discussed, as well as the need for further research to better understand the mechanisms of complement-mediated retinal damage in these disease states. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Effects of oakmoss and its components on biofilm formation of Legionella pneumophila.

PubMed

Nomura, Harue; Isshiki, Yasunori; Sakuda, Keisuke; Sakuma, Katsuya; Kondo, Seiichi

2013-01-01

Oakmoss and its components are known as antibacterial agents, specifically against Legionella pneumophila. In the present study, we investigated the effects of oakmoss and its components (phenol, didepside and isochromen derivatives) on L. pneumophila biofilm formation, with particular reference to the bactericidal activity (minimum bactericidal concentration; MBC) of these components against the bacterial cells in the biofilm. Of the 20 compounds tested, two didepside derivatives and four phenol derivatives reduced biofilm formation by more than 50% of that observed for the control at their respective minimum inhibitory concentrations (1/2×MIC). The inhibitory activities of these compounds were either equivalent to or greater than that of the clarithromycin reference. Isochromen derivatives had no effect on biofilm formation. Analysis of bactericidal activity of didepside and isochromen derivatives revealed that three of four didepside derivatives and one of four isochromen derivatives exhibited high bactericidal activity (MBC: 32.0-74.7 µg/mL) against the L. pneumophila in the biofilm after 24 h or 48 h of co-incubation; the antibacterial activities of these compounds were almost equivalent to clarithromycin and chlorhexidine gluconate (MBC: 42.7-64.0 µg/mL) that were used as references. Thus, based on their anti-biofilm forming and bactericidal activities, didepside derivatives are considered to be good candidates for disinfectants against L. pneumophila.

Recent advances in engineering topography mediated antibacterial surfaces

PubMed Central

Hasan, Jafar

2015-01-01

The tendency of bacterial cells to adhere and colonize a material surface leading to biofilm formation is a fundamental challenge underlying many different applications including microbial infections associated with biomedical devices and products. Although, bacterial attachment to surfaces has been extensively studied in the past, the effect of surface topography on bacteria–material interactions has received little attention until more recently. We review the recent progress in surface topography based approaches for engineering antibacterial surfaces. Biomimicry of antibacterial surfaces in nature is a popular strategy. Whereas earlier endeavors in the field aimed at minimizing cell attachment, more recent efforts have focused on developing bactericidal surfaces. However, not all such topography mediated bactericidal surfaces are necessarily cytocompatible thus underscoring the need for continued efforts for research in this area for developing antibacterial and yet cytocompatible surfaces for use in implantable biomedical applications. This mini-review provides a brief overview of the current strategies and challenges in the emerging field of topography mediated antibacterial surfaces. PMID:26372264

Recent advances in engineering topography mediated antibacterial surfaces

NASA Astrophysics Data System (ADS)

Hasan, Jafar; Chatterjee, Kaushik

2015-09-01

The tendency of bacterial cells to adhere and colonize a material surface leading to biofilm formation is a fundamental challenge underlying many different applications including microbial infections associated with biomedical devices and products. Although, bacterial attachment to surfaces has been extensively studied in the past, the effect of surface topography on bacteria-material interactions has received little attention until more recently. We review the recent progress in surface topography based approaches for engineering antibacterial surfaces. Biomimicry of antibacterial surfaces in nature is a popular strategy. Whereas earlier endeavors in the field aimed at minimizing cell attachment, more recent efforts have focused on developing bactericidal surfaces. However, not all such topography mediated bactericidal surfaces are necessarily cytocompatible thus underscoring the need for continued efforts for research in this area for developing antibacterial and yet cytocompatible surfaces for use in implantable biomedical applications. This mini-review provides a brief overview of the current strategies and challenges in the emerging field of topography mediated antibacterial surfaces.

Corneal epithelial wound healing and bactericidal effect of conditioned medium from human uterine cervical stem cells.

PubMed

Bermudez, Maria A; Sendon-Lago, Juan; Eiro, Noemi; Treviño, Mercedes; Gonzalez, Francisco; Yebra-Pimentel, Eva; Giraldez, Maria Jesus; Macia, Manuel; Lamelas, Maria Luz; Saa, Jorge; Vizoso, Francisco; Perez-Fernandez, Roman

2015-01-22

To evaluate the effect of conditioned medium from human uterine cervical stem cells (CM-hUCESCs) on corneal epithelial healing in a rat model of dry eye after alkaline corneal epithelial ulcer. We also tested the bactericidal effect of CM-hUCESCs. Dry eye was induced in rats by extraocular lacrimal gland excision, and corneal ulcers were produced using NaOH. Corneal histologic evaluation was made with hematoxylin-eosin (H&E) staining. Real-time PCR was used to evaluate mRNA expression levels of proinflammatory cytokines. We also studied the bactericidal effect of CM-hUCESCs in vitro and on infected corneal contact lenses (CLs) using Escherichia coli and Staphylococcus epidermidis bacteria. In addition, in order to investigate proteins from CM-hUCESCs that could mediate these effects, we carried out a human cytokine antibody array. After injury, dry eyes treated with CM-hUCESCs significantly improved epithelial regeneration and showed reduced corneal macrophage inflammatory protein-1 alpha (MIP-1α) and TNF-α mRNA expression as compared to untreated eyes and eyes treated with culture medium or sodium hyaluronate ophthalmic drops. In addition, we found in CM-hUCESCs high levels of proteins, such as tissue inhibitors of metalloproteinases 1 and 2, fibroblast growth factor 6 and 7, urokinase receptor, and hepatocyte growth factor, that could mediate these effects. In vitro, CM-hUCESCs showed a clear bactericidal effect on both E. coli and S. epidermidis and CLs infected with S. epidermidis. Analyses of CM-hUCESCs showed elevated levels of proteins that could be involved in the bactericidal effect, such as the chemokine (C-X-C motif) ligands 1, 6, 8, 10, and the chemokine (C-C motif) ligands 5 and 20. Treatment with CM-hUCESCs improved wound healing of alkali-injured corneas and showed a strong bactericidal effect on CLs. Patients using CLs and suffering from dry eye, allergies induced by commercial solutions, or small corneal injuries could benefit from this treatment. Copyright 2015 The Association for Research in Vision and Ophthalmology, Inc.

[Bactericidal activity of colloidal silver against grampositive and gramnegative bacteria].

PubMed

Afonina, I A; Kraeva, L A; Tseneva, G Ia

2010-01-01

It was shown that colloidal silver solution prepared in cooperation with the A. F. Ioffe Physical Technical Institute of the Russian Academy of Sciences, had significant bactericidal activity. Stable bactericidal effect on gramnegative microorganisms was observed after their 2-hour exposition in the solution of colloidal silver at a concentration of 10 ppm. Grampositive capsule-forming microorganisms were less susceptible to the colloidal silver solution: their death was observed after the 4-hour exposition in the solution.

Complement deposition in autoimmune hemolytic anemia is a footprint for difficult-to-detect IgM autoantibodies

PubMed Central

Meulenbroek, Elisabeth M.; de Haas, Masja; Brouwer, Conny; Folman, Claudia; Zeerleder, Sacha S.; Wouters, Diana

2015-01-01

In autoimmune hemolytic anemia autoantibodies against erythrocytes lead to increased clearance of the erythrocytes, which in turn results in a potentially fatal hemolytic anemia. Depending on whether IgG or IgM antibodies are involved, response to therapy is different. Proper identification of the isotype of the anti-erythrocyte autoantibodies is, therefore, crucial. However, detection of IgM autoantibodies can be challenging. We, therefore, set out to improve the detection of anti-erythrocyte IgM. Direct detection using a flow cytometry-based approach did not yield satisfactory improvements. Next, we analyzed whether the presence of complement C3 on a patient’s erythrocytes could be used for indirect detection of anti-erythrocyte IgM. To this end, we fractionated patients’ sera by size exclusion chromatography and tested which fractions yielded complement deposition on erythrocytes. Strikingly, we found that all patients with C3 on their erythrocytes according to standard diagnostic tests had an IgM anti-erythrocyte component that could activate complement, even if no such autoantibody had been detected with any other test. This also included all tested patients with only IgG and C3 on their erythrocytes, who would previously have been classified as having an IgG-only mediated autoimmune hemolytic anemia. Depleting patients’ sera of either IgG or IgM and testing the remaining complement activation confirmed this result. In conclusion, complement activation in autoimmune hemolytic anemia is mostly IgM-mediated and the presence of covalent C3 on patients’ erythrocytes can be taken as a footprint of the presence of anti-erythrocyte IgM. Based on this finding, we propose a diagnostic workflow that will aid in choosing the optimal treatment strategy. PMID:26354757

Trichinella spiralis Calreticulin Binds Human Complement C1q As an Immune Evasion Strategy.

PubMed

Zhao, Limei; Shao, Shuai; Chen, Yi; Sun, Ximeng; Sun, Ran; Huang, Jingjing; Zhan, Bin; Zhu, Xinping

2017-01-01

As a multicellular parasitic nematode, Trichinella spiralis regulates host immune responses by producing a variety of immunomodulatory molecules to escape from host immune attack, but the mechanisms underlying the immune evasion are not well understood. Here, we identified that T. spiralis calreticulin ( Ts -CRT), a Ca 2+ -binding protein, facilitated T. spiralis immune evasion by interacting with the first component of human classical complement pathway, C1q. In the present study, Ts -CRT was found to be expressed on the surface of different developmental stages of T. spiralis as well as in the secreted products of adult and muscle larval worms. Functional analysis identified that Ts -CRT was able to bind to human C1q, resulting in the inhibition of C1q-initiated complement classical activation pathway reflected by reduced C4/C3 generation and C1q-dependent lysis of antibody-sensitized sheep erythrocytes. Moreover, recombinant Ts -CRT (r Ts -CRT) binding to C1q suppressed C1q-induced THP-1-derived macrophages chemotaxis and reduced monocyte-macrophages release of reactive oxygen intermediates (ROIs). Blocking Ts -CRT on the surface of newborn larvae (NBL) of T. spiralis with anti- Ts -CRT antibody increased the C1q-mediated adherence of monocyte-macrophages to larvae and impaired larval infectivity. All of these results suggest that T. spiralis -expressed Ts -CRT plays crucial roles in T. spiralis immune evasion and survival in host mostly by directly binding to host complement C1q, which not only reduces C1q-mediated activation of classical complement pathway but also inhibits the C1q-induced non-complement activation of macrophages.

Trichinella spiralis Calreticulin Binds Human Complement C1q As an Immune Evasion Strategy

PubMed Central

Zhao, Limei; Shao, Shuai; Chen, Yi; Sun, Ximeng; Sun, Ran; Huang, Jingjing; Zhan, Bin; Zhu, Xinping

2017-01-01

As a multicellular parasitic nematode, Trichinella spiralis regulates host immune responses by producing a variety of immunomodulatory molecules to escape from host immune attack, but the mechanisms underlying the immune evasion are not well understood. Here, we identified that T. spiralis calreticulin (Ts-CRT), a Ca2+-binding protein, facilitated T. spiralis immune evasion by interacting with the first component of human classical complement pathway, C1q. In the present study, Ts-CRT was found to be expressed on the surface of different developmental stages of T. spiralis as well as in the secreted products of adult and muscle larval worms. Functional analysis identified that Ts-CRT was able to bind to human C1q, resulting in the inhibition of C1q-initiated complement classical activation pathway reflected by reduced C4/C3 generation and C1q-dependent lysis of antibody-sensitized sheep erythrocytes. Moreover, recombinant Ts-CRT (rTs-CRT) binding to C1q suppressed C1q-induced THP-1-derived macrophages chemotaxis and reduced monocyte–macrophages release of reactive oxygen intermediates (ROIs). Blocking Ts-CRT on the surface of newborn larvae (NBL) of T. spiralis with anti-Ts-CRT antibody increased the C1q-mediated adherence of monocyte–macrophages to larvae and impaired larval infectivity. All of these results suggest that T. spiralis-expressed Ts-CRT plays crucial roles in T. spiralis immune evasion and survival in host mostly by directly binding to host complement C1q, which not only reduces C1q-mediated activation of classical complement pathway but also inhibits the C1q-induced non-complement activation of macrophages. PMID:28620388

A De Novo Deletion in the Regulators of Complement Activation Cluster Producing a Hybrid Complement Factor H/Complement Factor H-Related 3 Gene in Atypical Hemolytic Uremic Syndrome.

PubMed

Challis, Rachel C; Araujo, Geisilaine S R; Wong, Edwin K S; Anderson, Holly E; Awan, Atif; Dorman, Anthony M; Waldron, Mary; Wilson, Valerie; Brocklebank, Vicky; Strain, Lisa; Morgan, B Paul; Harris, Claire L; Marchbank, Kevin J; Goodship, Timothy H J; Kavanagh, David

2016-06-01

The regulators of complement activation cluster at chromosome 1q32 contains the complement factor H (CFH) and five complement factor H-related (CFHR) genes. This area of the genome arose from several large genomic duplications, and these low-copy repeats can cause genome instability in this region. Genomic disorders affecting these genes have been described in atypical hemolytic uremic syndrome, arising commonly through nonallelic homologous recombination. We describe a novel CFH/CFHR3 hybrid gene secondary to a de novo 6.3-kb deletion that arose through microhomology-mediated end joining rather than nonallelic homologous recombination. We confirmed a transcript from this hybrid gene and showed a secreted protein product that lacks the recognition domain of factor H and exhibits impaired cell surface complement regulation. The fact that the formation of this hybrid gene arose as a de novo event suggests that this cluster is a dynamic area of the genome in which additional genomic disorders may arise. Copyright © 2016 by the American Society of Nephrology.

Natural IgM mediates complement-dependent uptake of Francisella tularensis by human neutrophils via CR1 and CR3 in nonimmune serum

PubMed Central

Schwartz, Justin T.; Barker, Jason H.; Long, Matthew E.; Kaufman, Justin; McCracken, Jenna; Allen, Lee-Ann H.

2012-01-01

A fundamental step in the life cycle of F. tularensis is bacterial entry into host cells. F. tularensis activates complement, and recent data suggest that the classical pathway is required for complement factor C3 deposition on the bacterial surface. Nevertheless, C3 deposition is inefficient and neither the specific serum components necessary for classical pathway activation by F. tularensis in nonimmune human serum, nor the receptors that mediate infection of neutrophils has been defined. Herein human neutrophil uptake of GFP-expressing F. tularensis strains LVS and Schu S4 was quantified with high efficiency by flow cytometry. Using depleted sera and purified complement components we demonstrated first that C1q and C3 were essential for F. tularensis phagocytosis whereas C5 was not. Second, we used purification and immuno-depletion approaches to identify a critical role for natural IgM in this process, and then used a wbtA2 mutant to identify LPS O-antigen and capsule as prominent targets of these antibodies on the bacterial surface. Finally, we demonstrate using receptor-blocking antibodies that CR1 (CD35) and CR3 (CD11b/CD18) acted in concert for phagocytosis of opsonized F. tularensis by human neutrophils, whereas CR3 and CR4 (CD11c/CD18) mediated infection of human monocyte-derived macrophages. Altogether, our data provide fundamental insight into mechanisms of F. tularensis phagocytosis and support a model whereby natural IgM binds to surface capsular and O-antigen polysaccharides of F. tularensis and initiates the classical complement cascade via C1q to promote C3-opsonization of the bacterium and phagocytosis via CR3 and either CR1 or CR4 in a phagocyte-specific manner. PMID:22888138

Polyhexamethylene guanidine hydrochloride shows bactericidal advantages over chlorhexidine digluconate against ESKAPE bacteria.

PubMed

Zhou, Zhongxin; Wei, Dafu; Lu, Yanhua

2015-01-01

More information regarding the bactericidal properties of polyhexamethylene guanidine hydrochloride (PHMG) against clinically important antibiotic-resistant ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) pathogens needs to be provided for its uses in infection control. The bactericidal properties of PHMG and chlorhexidine digluconate (CHG) were compared based on their minimum inhibitory concentrations (MICs), minimum bactericidal concentrations, and time-course-killing curves against clinically important antibiotic-susceptible and antibiotic-resistant ESKAPE pathogens. Results showed that PHMG exhibited significantly higher bactericidal activities against methicillin-resistant Staphylococcus aureus, carbapenem-resistant Klebsiella pneumoniae, and ceftazidime-resistant Enterobacter spp. than CHG. A slight bactericidal advantage over CHG was obtained against vancomycin-resistant Enterococcus faecium, ciprofloxacin- and levofloxacin-resistant Acinetobacter spp., and multidrug-resistant Pseudomonas aeruginosa. In previous reports, PHMG had higher antimicrobial activity against almost all tested Gram-negative bacteria and several Gram-positive bacteria than CHG using MIC test. These studies support the further development of covalently bound PHMG in sterile-surface materials and the incorporation of PHMG in novel disinfectant formulas. © 2014 International Union of Biochemistry and Molecular Biology, Inc.

Keeping It All Going-Complement Meets Metabolism.

PubMed

Kolev, Martin; Kemper, Claudia

2017-01-01

The complement system is an evolutionary old and crucial component of innate immunity, which is key to the detection and removal of invading pathogens. It was initially discovered as a liver-derived sentinel system circulating in serum, the lymph, and interstitial fluids that mediate the opsonization and lytic killing of bacteria, fungi, and viruses and the initiation of the general inflammatory responses. Although work performed specifically in the last five decades identified complement also as a critical instructor of adaptive immunity-indicating that complement's function is likely broader than initially anticipated-the dominant opinion among researchers and clinicians was that the key complement functions were in principle defined. However, there is now a growing realization that complement activity goes well beyond "classic" immune functions and that this system is also required for normal (neuronal) development and activity and general cell and tissue integrity and homeostasis. Furthermore, the recent discovery that complement activation is not confined to the extracellular space but occurs within cells led to the surprising understanding that complement is involved in the regulation of basic processes of the cell, particularly those of metabolic nature-mostly via novel crosstalks between complement and intracellular sensor, and effector, pathways that had been overlooked because of their spatial separation. These paradigm shifts in the field led to a renaissance in complement research and provide new platforms to now better understand the molecular pathways underlying the wide-reaching effects of complement functions in immunity and beyond. In this review, we will cover the current knowledge about complement's emerging relationship with the cellular metabolism machinery with a focus on the functional differences between serum-circulating versus intracellularly active complement during normal cell survival and induction of effector functions. We will also discuss how taking a closer look into the evolution of key complement components not only made the functional connection between complement and metabolism rather "predictable" but how it may also give clues for the discovery of additional roles for complement in basic cellular processes.

In vitro bactericidal and bacteriolytic activity of ceragenin CSA-13 against planktonic cultures and biofilms of Streptococcus pneumoniae and other pathogenic streptococci.

PubMed

Moscoso, Miriam; Esteban-Torres, María; Menéndez, Margarita; García, Ernesto

2014-01-01

Ceragenin CSA-13, a cationic steroid, is here reported to show a concentration-dependent bactericidal/bacteriolytic activity against pathogenic streptococci, including multidrug-resistant Streptococcus pneumoniae. The autolysis promoted by CSA-13 in pneumococcal cultures appears to be due to the triggering of the major S. pneumoniae autolysin LytA, an N-acetylmuramoyl-L-alanine amidase. CSA-13 also disintegrated pneumococcal biofilms in a very efficient manner, although at concentrations slightly higher than those required for bactericidal activity on planktonic bacteria. CSA-13 has little hemolytic activity which should allow testing its antibacterial efficacy in animal models.

The activity of several newer antimicrobials against logarithmically multiplying M. leprae in mice.

PubMed

Burgos, Jasmin; de la Cruz, Eduardo; Paredes, Rose; Andaya, Cora Revelyn; Gelber, Robert H

2011-09-01

Moxifloxacin, rifampicin, rifapentine, linezolid, and PA 824, alone and in combination, have been previously administered, as single doses and five times daily doses, to M. leprae infected mice during lag phase multiplication and were each found to have some bactericidal activity. The fluroquinolones, ofloxacin, moxifloxacin and gatifloxacin, (50 mg/kg, 150 mg/kg and 300 mg/kg) and the rifamycins (5 mg/kg, 10 mg/kg, and 20 mg/kg), rifampicin and rifapentine, were evaluated alone and in combination for bactericidal activity against M. leprae using the mouse footpad model during logarithmic multiplication. Linezolid and PA 824 were similarly evaluated alone and linezolid in combination with rifampicin, minocycline and ofloxacin. The three fluroquinolones and rifamycins were found alone and in combination to be bactericidal at all dosage schedules. PA 824 had no activity against M. leprae, while linezolid at a dose of 25 mg/kg was bacteriostatic, and progressively more bactericidal at doses of 50 mg/kg and 100 mg/kg. No antagonisms were detected between any of these drugs when used in combinations. Moxifloxacin, gatifloxacin, rifapentine and linezolid were found bactericidal against rapidly multiplying M. leprae.

Novel roles of complement in renal diseases and their therapeutic consequences.

PubMed

Wada, Takehiko; Nangaku, Masaomi

2013-09-01

The complement system functions as a part of the innate immune system. Inappropriate activation of the complement pathways has a deleterious effect on kidneys. Recent advances in complement research have provided new insights into the pathogenesis of glomerular and tubulointerstitial injury associated with complement activation. A new disease entity termed 'C3 glomerulopathy' has recently been proposed and is characterized by isolated C3 deposition in glomeruli without positive staining for immunoglobulins. Genetic and functional studies have demonstrated that several different mutations and disease variants, as well as the generation of autoantibodies, are potentially associated with its pathogenesis. The data from comprehensive analyses suggest that complement dysregulation can also be associated with hemolytic uremic syndrome and more common glomerular diseases, such as IgA nephropathy and diabetic kidney disease. In addition, animal studies utilizing genetically modified mice have begun to elucidate the molecular pathomechanisms associated with the complement system. From a diagnostic point of view, a noninvasive, MRI-based method for detecting C3 has recently been developed to serve as a novel tool for diagnosing complement-mediated kidney diseases. While novel therapeutic tools related to complement regulation are emerging, studies evaluating the precise roles of the complement system in kidney diseases will still be useful for developing new therapeutic approaches.

Bactericidal/permeability-increasing protein in the reproductive system of male mice may be involved in the sperm-oocyte fusion.

PubMed

Li, Kun; Liu, Yue; Xia, Xiaoyu; Wang, Li; Lu, Meige; Hu, Yanqin; Xu, Chen

2013-08-01

Bactericidal/permeability-increasing protein (BPI) is a 455-residue (∼55 kDa) protein found mainly in the primary (azurophilic) granules of human neutrophils. BPI is an endogenous antibiotic protein that belongs to the family of mammalian lipopolysaccharide (LPS)-binding and lipid transport proteins. Its major function is to kill Gram-negative bacteria, thereby protecting the host from infection. In addition, BPI can inhibit angiogenesis, suppress LPS-mediated platelet activation, increase DNA synthesis, and activate ERK/Akt signaling. In this study, we found that Bpi was expressed in the testis and epididymis but not in the seminal vesicles, prostate, and solidification glands. BPI expression in the epididymis increased upon upregulation of testosterone, caused by injection of GNRH. In orchidectomized mice, BPI expression was significantly reduced, but its expression was restored to 30% of control levels in orchidectomized mice that received supplementary testosterone. The number of sperm fused per egg significantly decreased after incubation with anti-BPI antiserum. These results suggest that BPI may take part in the process of sperm-oocyte fusion and play a unique and significant role in reproduction.

Catalase Expression Is Modulated by Vancomycin and Ciprofloxacin and Influences the Formation of Free Radicals in Staphylococcus aureus Cultures

PubMed Central

Wang, Ying; Hougaard, Anni B.; Paulander, Wilhelm; Skibsted, Leif H.

2015-01-01

Detection of free radicals in biological systems is challenging due to their short half-lives. We have applied electron spin resonance (ESR) spectroscopy combined with spin traps using the probes PBN (N-tert-butyl-α-phenylnitrone) and DMPO (5,5-dimethyl-1-pyrroline N-oxide) to assess free radical formation in the human pathogen Staphylococcus aureus treated with a bactericidal antibiotic, vancomycin or ciprofloxacin. While we were unable to detect ESR signals in bacterial cells, hydroxyl radicals were observed in the supernatant of bacterial cell cultures. Surprisingly, the strongest signal was detected in broth medium without bacterial cells present and it was mitigated by iron chelation or by addition of catalase, which catalyzes the decomposition of hydrogen peroxide to water and oxygen. This suggests that the signal originates from hydroxyl radicals formed by the Fenton reaction, in which iron is oxidized by hydrogen peroxide. Previously, hydroxyl radicals have been proposed to be generated within bacterial cells in response to bactericidal antibiotics. We found that when S. aureus was exposed to vancomycin or ciprofloxacin, hydroxyl radical formation in the broth was indeed increased compared to the level seen with untreated bacterial cells. However, S. aureus cells express catalase, and the antibiotic-mediated increase in hydroxyl radical formation was correlated with reduced katA expression and catalase activity in the presence of either antibiotic. Therefore, our results show that in S. aureus, bactericidal antibiotics modulate catalase expression, which in turn influences the formation of free radicals in the surrounding broth medium. If similar regulation is found in other bacterial species, it might explain why bactericidal antibiotics are perceived as inducing formation of free radicals. PMID:26150471

Catalase Expression Is Modulated by Vancomycin and Ciprofloxacin and Influences the Formation of Free Radicals in Staphylococcus aureus Cultures.

PubMed

Wang, Ying; Hougaard, Anni B; Paulander, Wilhelm; Skibsted, Leif H; Ingmer, Hanne; Andersen, Mogens L

2015-09-01

Detection of free radicals in biological systems is challenging due to their short half-lives. We have applied electron spin resonance (ESR) spectroscopy combined with spin traps using the probes PBN (N-tert-butyl-α-phenylnitrone) and DMPO (5,5-dimethyl-1-pyrroline N-oxide) to assess free radical formation in the human pathogen Staphylococcus aureus treated with a bactericidal antibiotic, vancomycin or ciprofloxacin. While we were unable to detect ESR signals in bacterial cells, hydroxyl radicals were observed in the supernatant of bacterial cell cultures. Surprisingly, the strongest signal was detected in broth medium without bacterial cells present and it was mitigated by iron chelation or by addition of catalase, which catalyzes the decomposition of hydrogen peroxide to water and oxygen. This suggests that the signal originates from hydroxyl radicals formed by the Fenton reaction, in which iron is oxidized by hydrogen peroxide. Previously, hydroxyl radicals have been proposed to be generated within bacterial cells in response to bactericidal antibiotics. We found that when S. aureus was exposed to vancomycin or ciprofloxacin, hydroxyl radical formation in the broth was indeed increased compared to the level seen with untreated bacterial cells. However, S. aureus cells express catalase, and the antibiotic-mediated increase in hydroxyl radical formation was correlated with reduced katA expression and catalase activity in the presence of either antibiotic. Therefore, our results show that in S. aureus, bactericidal antibiotics modulate catalase expression, which in turn influences the formation of free radicals in the surrounding broth medium. If similar regulation is found in other bacterial species, it might explain why bactericidal antibiotics are perceived as inducing formation of free radicals. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

In vitro bactericidal activity of enrofloxacin against gyrA mutant and qnr-containing Escherichia coli isolates from animals.

PubMed

Cengiz, M; Sahinturk, P; Sonal, S; Buyukcangaz, E; Sen, A; Arslan, E

2013-05-04

The objective of this work was to investigate the bactericidal activity of enrofloxacin against gyrA mutant and qnr-containing Escherichia coli isolates from animals. The minimum inhibitory concentrations (MICs) of gyrA mutant and qnr-containing E coli isolates ranged from 1 µg/ml to 32 µg/ml for enrofloxacin. Time-kill experiments were performed using selected E coli isolates. For the time-kill experiments, the colony counts were determined by plating each diluted sample onto plate count agar and an integrated pharmacokinetic/pharmacodynamics area measure (log ratio area) was applied to the colony-forming units (cfu) data. In general, enrofloxacin exhibited bactericidal activity against all the gyrA mutant E coli isolates at all concentrations greater than four times the MIC. However, the bactericidal activity of enrofloxacin for all the qnr-containing E coli isolates was less dependent on concentration. The results of the present study indicated that the genetic mechanism of resistance might account for the different bactericidal activities of enrofloxacin observed for the gyrA mutant and the qnr-containing E coli isolates. Therefore, in addition to MIC assays, genetic mechanism-based pharmacodynamic models should be used to provide accurate predictions of the effects of drugs on resistant bacteria.

The Effect of Long-Term Storage on the Physiochemical and Bactericidal Properties of Electrochemically Activated Solutions

PubMed Central

Robinson, Gareth; Thorn, Robin; Reynolds, Darren

2013-01-01

Electrochemically activated solutions (ECAS) are generated by electrolysis of NaCl solutions, and demonstrate broad spectrum antimicrobial activity and high environmental compatibility. The biocidal efficacy of ECAS at the point of production is widely reported in the literature, as are its credentials as a “green biocide.” Acidic ECAS are considered most effective as biocides at the point of production and ill suited for extended storage. Acidic ECAS samples were stored at 4 °C and 20 °C in glass and polystyrene containers for 398 days, and tested for free chlorine, pH, ORP and bactericidal activity throughout. ORP and free chlorine (mg/L) in stored ECAS declined over time, declining at the fastest rate when stored at 20 °C in polystyrene and at the slowest rate when stored at 4 °C in glass. Bactericidal efficacy was also affected by storage and ECAS failed to produce a 5 log10 reduction on five occasions when stored at 20 °C. pH remained stable throughout the storage period. This study represents the longest storage evaluation of the physiochemical parameters and bactericidal efficacy of acidic ECAS within the published literature and reveals that acidic ECAS retain useful bactericidal activity for in excess of 12 months, widening potential applications. PMID:23263673

The effect of long-term storage on the physiochemical and bactericidal properties of electrochemically activated solutions.

PubMed

Robinson, Gareth; Thorn, Robin; Reynolds, Darren

2012-12-24

Electrochemically activated solutions (ECAS) are generated by electrolysis of NaCl solutions, and demonstrate broad spectrum antimicrobial activity and high environmental compatibility. The biocidal efficacy of ECAS at the point of production is widely reported in the literature, as are its credentials as a "green biocide." Acidic ECAS are considered most effective as biocides at the point of production and ill suited for extended storage. Acidic ECAS samples were stored at 4 °C and 20 °C in glass and polystyrene containers for 398 days, and tested for free chlorine, pH, ORP and bactericidal activity throughout. ORP and free chlorine (mg/L) in stored ECAS declined over time, declining at the fastest rate when stored at 20 °C in polystyrene and at the slowest rate when stored at 4 °C in glass. Bactericidal efficacy was also affected by storage and ECAS failed to produce a 5 log(10) reduction on five occasions when stored at 20 °C. pH remained stable throughout the storage period. This study represents the longest storage evaluation of the physiochemical parameters and bactericidal efficacy of acidic ECAS within the published literature and reveals that acidic ECAS retain useful bactericidal activity for in excess of 12 months, widening potential applications.

Suppression of bactericidal activity of human polymorphonuclear leukocytes by Bacteroides gingivalis.

PubMed Central

Yoneda, M; Maeda, K; Aono, M

1990-01-01

The direct effects of the culture supernatant of oral microorganisms on the bactericidal activity of human polymorphonuclear leukocytes (PMNs) were investigated. The bactericidal activity of PMNs, which were preincubated with the supernatant of Bacteroides gingivalis, Bacteroides intermedius, Bacteroides melaninogenicus or phosphate-buffered saline, was examined by counting the surviving bacteria. B. gingivalis-treated PMNs were found to have a diminished ability for killing bacteria in the presence or absence of serum. The chemiluminescence response of PMNs, which were preincubated with the culture supernatant of B. gingivalis, was strikingly reduced compared with that of PMNs preincubated with phosphate-buffered saline or other bacterial culture supernatants. The production of superoxide anion (O2-) by PMNs stimulated with either formyl-methionyl-leucyl-phenylalanine or phorbol myristate acetate was reduced in both cases after the PMNs were preincubated with the culture supernatant of B. gingivalis. However, it was observed that there was more reduction in superoxide anion (O2-) production stimulated with formyl-methionyl-leucyl-phenylalanine compared with that stimulated with phorbol myristate acetate. These results suggest that B. gingivalis releases a factor which interferes with the bactericidal activity of PMNs by modulating the generation of reactive oxygen species. These suppressive effects on bactericidal activity may be important in the pathogenesis of this microorganism. PMID:2153632

Multi-functional mechanisms of immune evasion by the streptococcal complement inhibitor C5a peptidase

PubMed Central

Reglinski, Mark; Calay, Damien; Siggins, Matthew K.; Mason, Justin C.; Botto, Marina; Sriskandan, Shiranee

2017-01-01

The complement cascade is crucial for clearance and control of invading pathogens, and as such is a key target for pathogen mediated host modulation. C3 is the central molecule of the complement cascade, and plays a vital role in opsonization of bacteria and recruitment of neutrophils to the site of infection. Streptococcal species have evolved multiple mechanisms to disrupt complement-mediated innate immunity, among which ScpA (C5a peptidase), a C5a inactivating enzyme, is widely conserved. Here we demonstrate for the first time that pyogenic streptococcal species are capable of cleaving C3, and identify C3 and C3a as novel substrates for the streptococcal ScpA, which are functionally inactivated as a result of cleavage 7 amino acids upstream of the natural C3 convertase. Cleavage of C3a by ScpA resulted in disruption of human neutrophil activation, phagocytosis and chemotaxis, while cleavage of C3 generated abnormally-sized C3a and C3b moieties with impaired function, in particular reducing C3 deposition on the bacterial surface. Despite clear effects on human complement, expression of ScpA reduced clearance of group A streptococci in vivo in wildtype and C5 deficient mice, and promoted systemic bacterial dissemination in mice that lacked both C3 and C5, suggesting an additional complement-independent role for ScpA in streptococcal pathogenesis. ScpA was shown to mediate streptococcal adhesion to both human epithelial and endothelial cells, consistent with a role in promoting bacterial invasion within the host. Taken together, these data show that ScpA is a multi-functional virulence factor with both complement-dependent and independent roles in streptococcal pathogenesis. PMID:28806402

Bactericidal Effects and Mechanism of Action of Olanexidine Gluconate, a New Antiseptic

PubMed Central

Iwata, Koushi; Nii, Takuya; Nakata, Hikaru; Tsubotani, Yoshie; Inoue, Yasuhide

2015-01-01

Olanexidine gluconate [1-(3,4-dichlorobenzyl)-5-octylbiguanide gluconate] (development code OPB-2045G) is a new monobiguanide compound with bactericidal activity. In this study, we assessed its spectrum of bactericidal activity and mechanism of action. The minimal bactericidal concentrations of the compound for 30-, 60-, and 180-s exposures were determined with the microdilution method using a neutralizer against 320 bacterial strains from culture collections and clinical isolates. Based on the results, the estimated bactericidal olanexidine concentrations with 180-s exposures were 869 μg/ml for Gram-positive cocci (155 strains), 109 μg/ml for Gram-positive bacilli (29 strains), and 434 μg/ml for Gram-negative bacteria (136 strains). Olanexidine was active against a wide range of bacteria, especially Gram-positive cocci, including methicillin-resistant Staphylococcus aureus and vancomycin-resistant enterococci, and had a spectrum of bactericidal activity comparable to that of commercial antiseptics, such as chlorhexidine and povidone-iodine. In vitro experiments exploring its mechanism of action indicated that olanexidine (i) interacts with the bacterial surface molecules, such as lipopolysaccharide and lipoteichoic acid, (ii) disrupts the cell membranes of liposomes, which are artificial bacterial membrane models, (iii) enhances the membrane permeability of Escherichia coli, (iv) disrupts the membrane integrity of S. aureus, and (v) denatures proteins at relatively high concentrations (≥160 μg/ml). These results indicate that olanexidine probably binds to the cell membrane, disrupts membrane integrity, and its bacteriostatic and bactericidal effects are caused by irreversible leakage of intracellular components. At relatively high concentrations, olanexidine aggregates cells by denaturing proteins. This mechanism differs slightly from that of a similar biguanide compound, chlorhexidine. PMID:25987609

[Antibacterial actin of vinegar against food-borne pathogenic bacteria including Escherichia coli O157:H7 (Part 2). Effect of sodium chloride and temperature on bactericidal activity].

PubMed

Entani, E; Asai, M; Tsujihata, S; Tsukamoto, Y; Ohta, M

1997-05-01

Bactericidal effects of various kinds of AWASEZU (processed vinegar, 2.5% acidity) on food-borne pathogenic bacteria including Escherichia coli O157:H7 and other bacteria were examined. the order of bactericidal activities was NIHAIZU (3.5% NaCl was added) > SANBA-IZU (3.5% NaCl and 10% sucrose were added) > plain vinegar (spirit vinegar) > AMAZU (10% sucrose was added). This indicates that their activities were enhanced by the addition of sodium chloride and suppressed by the addition of sugar. On the other hand, when soy sauce was used instead of sodium chloride, the order of bactericidal activities was plain vinegar > AMAZU > NIHAIZU > SANBAIZU. This is mainly because their activities were suppressed by the increase in the pH value. The effect of sodium chloride (0.01-15%) and temperature (10-50 degrees C) on bactericidal activities against E. coli O157:H7 in spirit vinegar (0.5-2.5% acidity) was further examined. When vinegar was used in combination with sodium chloride, predominant synergism on the bactericidal activity was observed. Their activities were markedly enhanced by the addition of sodium chloride in proportion to the concentration. In addition to this, at higher temperatures spirit vinegar killed bacteria much more rapidly. It should be noted that the bactericidal activity of spirit vinegar was extremely enhanced by the combined use of the addition of sodium chloride and the rise of temperature. For example, in 2.5% acidity vinegar, the time required for 3 log decrease in viable cell numbers at 20 degrees C was shortened to 1/140-fold by the addition of 5% sodium chloride, shortened to 1/51-fold by the rise of the reaction temperature at 40 degrees C, and shortened to 1/830-fold; 0.89 minutes by both the addition of 5% sodium chloride and the rise of temperature at 40 degrees C. In order to propose the methods to prevent food poisoning by bacterial infection, bactericidal activities of vinegar solution containing sodium chloride on cooking tools and raw vegetables were examined. Vinegar solution (1-2% acidity, 3-7% NaCl) produced more than 3 log decrease in viable cell numbers of E. coli O157:H7 on the surface of cutting board, and cabbage and cucumber at 20-50 degrees C. These results suggested that the treatment with vinegar solution containing sodium chloride may be one of the useful methods to prevent food poisoning.

Smoke Exposure Causes Endoplasmic Reticulum Stress and Lipid Accumulation in Retinal Pigment Epithelium through Oxidative Stress and Complement Activation*

PubMed Central

Kunchithapautham, Kannan; Atkinson, Carl; Rohrer, Bärbel

2014-01-01

Age-related macular degeneration (AMD) is a complex disease caused by genetic and environmental factors, including genetic variants in complement components and smoking. Smoke exposure leads to oxidative stress, complement activation, endoplasmic reticulum (ER) stress, and lipid dysregulation, which have all been proposed to be associated with AMD pathogenesis. Here we examine the effects of smoke exposure on the retinal pigment epithelium (RPE). Mice were exposed to cigarette smoke or filtered air for 6 months. RPE cells grown as stable monolayers were exposed to 5% cigarette smoke extract (CSE). Effects of smoke were determined by biochemical, molecular, and histological measures. Effects of the alternative pathway (AP) of complement and complement C3a anaphylatoxin receptor signaling were analyzed using knock-out mice or specific inhibitors. ER stress markers were elevated after smoke exposure in RPE of intact mice, which was eliminated in AP-deficient mice. To examine this relationship further, RPE monolayers were exposed to CSE. Short term smoke exposure resulted in production and release of complement C3, the generation of C3a, oxidative stress, complement activation on the cell membrane, and ER stress. Long term exposure to CSE resulted in lipid accumulation, and secretion. All measures were reversed by blocking C3a complement receptor (C3aR), alternative complement pathway signaling, and antioxidant therapy. Taken together, our results provide clear evidence that smoke exposure results in oxidative stress and complement activation via the AP, resulting in ER stress-mediated lipid accumulation, and further suggesting that oxidative stress and complement act synergistically in the pathogenesis of AMD. PMID:24711457

Bactericidal activity of tracheal antimicrobial peptide against respiratory pathogens of cattle.

PubMed

Taha-Abdelaziz, Khaled; Perez-Casal, José; Schott, Courtney; Hsiao, Jason; Attah-Poku, Samuel; Slavić, Durđa; Caswell, Jeff L

2013-04-15

Tracheal antimicrobial peptide (TAP) is a β-defensin produced by mucosal epithelial cells of cattle. Although effective against several human pathogens, the activity of this bovine peptide against the bacterial pathogens that cause bovine respiratory disease have not been reported. This study compared the antibacterial effects of synthetic TAP against Mannheimia haemolytica, Histophilus somni, Pasteurella multocida, and Mycoplasma bovis. Bactericidal activity against M. bovis was not detected. In contrast, the Pasteurellaceae bacteria showed similar levels of susceptibility to that of Escherichia coli, with 0.125μg TAP inhibiting growth in a radial diffusion assay and minimum inhibitory concentrations of 1.56-6.25μg/ml in a bactericidal assay. Significant differences among isolates were not observed. Sequencing of exon 2 of the TAP gene from 23 cattle revealed a prevalent non-synonymous single nucleotide polymorphism (SNP) A137G, encoding either serine or asparagine at residue 20 of the mature peptide. The functional effect of this SNP was tested against M. haemolytica using synthetic peptides. The bactericidal effect of the asparagine-containing peptide was consistently higher than the serine-containing peptide. Bactericidal activities were similar for an acapsular mutant of M. haemolytica compared to the wild type. These findings indicate that the Pasteurellaceae bacteria that cause bovine respiratory disease are susceptible to killing by bovine TAP and appear not to have evolved resistance, whereas M. bovis appears to be resistant. A non-synonymous SNP was identified in the coding region of the TAP gene, and the corresponding peptides vary in their bactericidal activity against M. haemolytica. Copyright © 2013 Elsevier B.V. All rights reserved.

NF-κB and enhancer-binding CREB protein scaffolded by CREB-binding protein (CBP)/p300 proteins regulate CD59 protein expression to protect cells from complement attack.

PubMed

Du, Yiqun; Teng, Xiaoyan; Wang, Na; Zhang, Xin; Chen, Jianfeng; Ding, Peipei; Qiao, Qian; Wang, Qingkai; Zhang, Long; Yang, Chaoqun; Yang, Zhangmin; Chu, Yiwei; Du, Xiang; Zhou, Xuhui; Hu, Weiguo

2014-01-31

The complement system can be activated spontaneously for immune surveillance or induced to clear invading pathogens, in which the membrane attack complex (MAC, C5b-9) plays a critical role. CD59 is the sole membrane complement regulatory protein (mCRP) that restricts MAC assembly. CD59, therefore, protects innocent host cells from attacks by the complement system, and host cells require the constitutive and inducible expression of CD59 to protect themselves from deleterious destruction by complement. However, the mechanisms that underlie CD59 regulation remain largely unknown. In this study we demonstrate that the widely expressed transcription factor Sp1 may regulate the constitutive expression of CD59, whereas CREB-binding protein (CBP)/p300 bridge NF-κB and CREB, which surprisingly functions as an enhancer-binding protein to induce the up-regulation of CD59 during in lipopolysaccharide (LPS)-triggered complement activation, thus conferring host defense against further MAC-mediated destruction. Moreover, individual treatment with LPS, TNF-α, and the complement activation products (sublytic MAC (SC5b-9) and C5a) could increase the expression of CD59 mainly by activating NF-κB and CREB signaling pathways. Together, our findings identify a novel gene regulation mechanism involving CBP/p300, NF-κB, and CREB; this mechanism suggests potential drug targets for controlling various complement-related human diseases.

Structure of C3b reveals conformational changes that underlie complement activity.

PubMed

Janssen, Bert J C; Christodoulidou, Agni; McCarthy, Andrew; Lambris, John D; Gros, Piet

2006-11-09

Resistance to infection and clearance of cell debris in mammals depend on the activation of the complement system, which is an important component of innate and adaptive immunity. Central to the complement system is the activated form of C3, called C3b, which attaches covalently to target surfaces to amplify complement response, label cells for phagocytosis and stimulate the adaptive immune response. C3b consists of 1,560 amino-acid residues and has 12 domains. It binds various proteins and receptors to effect its functions. However, it is not known how C3 changes its conformation into C3b and thereby exposes its many binding sites. Here we present the crystal structure at 4-A resolution of the activated complement protein C3b and describe the conformational rearrangements of the 12 domains that take place upon proteolytic activation. In the activated form the thioester is fully exposed for covalent attachment to target surfaces and is more than 85 A away from the buried site in native C3 (ref. 5). Marked domain rearrangements in the alpha-chain present an altered molecular surface, exposing hidden and cryptic sites that are consistent with known putative binding sites of factor B and several complement regulators. The structural data indicate that the large conformational changes in the proteolytic activation and regulation of C3 take place mainly in the first conversion step, from C3 to C3b. These insights are important for the development of strategies to treat immune disorders that involve complement-mediated inflammation.

The Many Facets of Lipooligosaccharide as a Virulence Factor for Histophilus somni.

PubMed

Inzana, Thomas J

2016-01-01

The lipooligosaccharide (LOS) of Histophilus somni is a multifaceted molecule that provides critical protection to the bacterium against host defenses, may act as an adhesin, and like similar molecules of gram-negative bacteria, is an endotoxin that signals through toll-like receptor 4 and NF-κB to cause inflammation. The lipid A component is responsible for the endotoxic and apoptotic activity of the LOS. The H. somni LOS lacks O-side chains typically characteristic of gram-negative bacteria that have lipopolysaccharide, but has a complex, microheterogeneous outer core. The LOS of disease isolates is capable of undergoing structural and antigenic phase variation of its outer core due to slip-strand mispairing of glycosyltransferase genes that contain repetitive sequences of DNA base pairs. Such variation enables the bacteria to evade bactericidal antibodies made to oligosaccharide antigens. In addition, the LOS can be decorated with phase-variable phosphorylcholine (ChoP), which binds to platelet-activating factor receptor on host cells, thereby aiding in colonization of the upper respiratory tract. However, ChoP is likely not expressed when the bacteria are in systemic sites because ChoP also binds to C-reactive protein, resulting in activation of host complement and promoting bactericidal activity. The structure of some LOS outer core chains is identical to oligosaccharides on host glycosphingolipids of red blood cells, other cells, and merconium (lacto-N-neotetraose, lacto-N-biose, N-acetyllactosamine, etc.). Furthermore, terminal galactose residues on LOS and elsewhere are decorated with sialic acid, which blocks antibody binding, activation of complement, phagocytosis, and intracellular killing. Therefore, antigenic mimicry of host antigens is an important defense mechanism provided by the oligosaccharide component of the LOS to avoid innate and adaptive host defense mechanisms. However, some strains of H. somni isolated from the bovine genital tract, particularly the normal bovine prepuce, are incapable of LOS phase variation, sialylation of the LOS, and expression of ChoP. At least 1 such strain has been shown to be avirulent, underscoring the importance of the LOS as a virulence factor, although this strain is deficient in other factors as well. The structure and arrangement of the inner core glycoses (heptose and 3-deoxy-D-manno-2-octulosnic acid) is remarkably similar to the inner core oligosaccharide on some strains of Neisseria spp., and mutants that contain a truncated LOS oligosaccharide are considerably more serum-sensitive than the parent strain. Therefore, the LOS is a critical component that enables H. somni to resist host defenses and cause disease.

Antimicrobial properties of single-donor-derived, platelet-leukocyte fibrin for fistula occlusion: An in vitro study.

PubMed

Wu, Xiuwen; Ren, Jianan; Yuan, Yujie; Luan, Jianfeng; Yao, Genhong; Li, Jieshou

2013-01-01

Fibrin glue is a promising alternative for low-output enterocutaneous fistula closure. Bacterial flora colonizing inside the fistula tract, however, may limit the glue application. Single-donor-derived, platelet-rich materials were hypothesized in this study to have antimicrobial activity against Gram-negative microorganisms. Platelet-leukocyte fibrin (PLF), platelet-rich plasma (PRP), and platelet-poor plasma (PPP) were obtained from healthy volunteers. The amounts of platelet, leukocyte, and complement/antibody were determined. In vitro laboratory susceptibility to PLF and plasmas was determined by the Kirby-Bauer disc-diffusion method. Antimicrobial activity of PLF, PRP, and PPP against three Gram-negative ATCC strains was determined in a bacterial kill assay. Levels of complement and antibody did not significantly differ among PLF, PRP, and PPP (p > 0.05), while platelet and leukocyte counts in platelet-rich biomaterials were significantly higher than those in PPP (p < 0.001). The bactericidal effects produced by PLF and PRP were more pronounced than those by PPP in both qualitative and quantitative aspects. No inhibitory zones against three Gram-negative organisms were observed in the Bioseal®. In conclusion, our homemade PLF, as compared with commercial products, had remarkable antimicrobial activity against Gram-negative bacteria relevant to fistula colonization. The presence of platelets and leukocytes may play an important role in bacterial defense. This is the first study to demonstrate the antibacterial properties of single-unit PLF for fistula closure, presenting a new opportunity for glue sealing.

Differential Effects of Complement Activation Products C3a and C5a on Cardiovascular Function in Hypertensive Pregnant Rats

PubMed Central

Lillegard, Kathryn E.; Loeks-Johnson, Alex C.; Opacich, Jonathan W.; Peterson, Jenna M.; Bauer, Ashley J.; Elmquist, Barbara J.; Regal, Ronald R.; Gilbert, Jeffrey S.

2014-01-01

Early-onset pre-eclampsia is characterized by decreased placental perfusion, new-onset hypertension, angiogenic imbalance, and endothelial dysfunction associated with excessive activation of the innate immune complement system. Although our previous studies demonstrated that inhibition of complement activation attenuates placental ischemia–induced hypertension using the rat reduced uterine perfusion pressure (RUPP) model, the important product(s) of complement activation has yet to be identified. We hypothesized that antagonism of receptors for complement activation products C3a and C5a would improve vascular function and attenuate RUPP hypertension. On gestational day (GD) 14, rats underwent sham surgery or vascular clip placement on ovarian arteries and abdominal aorta (RUPP). Rats were treated once daily with the C5a receptor antagonist (C5aRA), PMX51 (acetyl-F-[Orn-P-(D-Cha)-WR]), the C3a receptor antagonist (C3aRA), SB290157 (N2-[(2,2-diphenylethoxy)acetyl]-l-arginine), or vehicle from GD 14–18. Both the C3aRA and C5aRA attenuated placental ischemia–induced hypertension without affecting the decreased fetal weight or decreased concentration of free circulating vascular endothelial growth factor (VEGF) also present in this model. The C5aRA, but not the C3aRA, attenuated placental ischemia–induced increase in heart rate and impaired endothelial-dependent relaxation. The C3aRA abrogated the acute pressor response to C3a peptide injection, but it also unexpectedly attenuated the placental ischemia–induced increase in C3a, suggesting nonreceptor-mediated effects. Overall, these results indicate that both C3a and C5a are important products of complement activation that mediate the hypertension regardless of the reduction in free plasma VEGF. The mechanism by which C3a contributes to placental ischemia–induced hypertension appears to be distinct from that of C5a, and management of pregnancy-induced hypertension is likely to require a broad anti-inflammatory approach. PMID:25150279

Site-targeted complement inhibition by a complement receptor 2-conjugated inhibitor (mTT30) ameliorates post-injury neuropathology in mouse brains.

PubMed

Rich, Megan C; Keene, Chesleigh N; Neher, Miriam D; Johnson, Krista; Yu, Zhao-Xue; Ganivet, Antoine; Holers, V Michael; Stahel, Philip F

2016-03-23

Intracerebral complement activation after severe traumatic brain injury (TBI) leads to a cascade of neuroinflammatory pathological sequelae that propagate host-mediated secondary brain injury and adverse outcomes. There are currently no specific pharmacological agents on the market to prevent or mitigate the development of secondary cerebral insults after TBI. A novel chimeric CR2-fH compound (mTT30) provides targeted inhibition of the alternative complement pathway at the site of tissue injury. This experimental study was designed to test the neuroprotective effects of mTT30 in a mouse model of closed head injury. The administration of 500 μg mTT30 i.v. at 1 h, 4 h and 24 h after head injury attenuated complement C3 deposition in injured brains, reduced the extent of neuronal cell death, and decreased post-injury microglial activation, compared to vehicle-injected placebo controls. These data imply that site-targeted alternative pathway complement inhibition may represent a new promising therapeutic avenue for the future management of severe TBI. Copyright © 2016. Published by Elsevier Ireland Ltd.

Evaluation of the Bactericidal Activity of Plazomicin and Comparators against Multidrug-resistant Enterobacteriaceae.

PubMed

Thwaites, M; Hall, D; Shinabarger, D; Serio, A W; Krause, K M; Marra, A; Pillar, C

2018-06-04

The next-generation aminoglycoside plazomicin, in development for infections due to multi-drug resistant (MDR) Enterobacteriaceae, was evaluated alongside comparators for bactericidal activity in minimum bactericidal concentration (MBC) and time-kill (TK) assays against MDR Enterobacteriaceae isolates with characterized aminoglycoside and β-lactam resistance mechanisms. Overall, plazomicin and colistin were the most potent, with plazomicin demonstrating an MBC 50/90 of 0.5/4 μg/mL and sustained 3-log 10 kill against MDR Escherichia coli , Klebsiella pneumoniae and Enterobacter spp. Copyright © 2018 Thwaites et al.

Bactericidal activity of biomimetic diamond nanocone surfaces.

PubMed

Fisher, Leanne E; Yang, Yang; Yuen, Muk-Fung; Zhang, Wenjun; Nobbs, Angela H; Su, Bo

2016-03-17

The formation of biofilms on implant surfaces and the subsequent development of medical device-associated infections are difficult to resolve and can cause considerable morbidity to the patient. Over the past decade, there has been growing recognition that physical cues, such as surface topography, can regulate biological responses and possess bactericidal activity. In this study, diamond nanocone-patterned surfaces, representing biomimetic analogs of the naturally bactericidal cicada fly wing, were fabricated using microwave plasma chemical vapor deposition, followed by bias-assisted reactive ion etching. Two structurally distinct nanocone surfaces were produced, characterized, and the bactericidal ability examined. The sharp diamond nanocone features were found to have bactericidal capabilities with the surface possessing the more varying cone dimension, nonuniform array, and decreased density, showing enhanced bactericidal ability over the more uniform, highly dense nanocone surface. Future research will focus on using the fabrication process to tailor surface nanotopographies on clinically relevant materials that promote both effective killing of a broader range of microorganisms and the desired mammalian cell response. This study serves to introduce a technology that may launch a new and innovative direction in the design of biomaterials with capacity to reduce the risk of medical device-associated infections.

Complement Evasion by Pathogenic Leptospira.

PubMed

Fraga, Tatiana Rodrigues; Isaac, Lourdes; Barbosa, Angela Silva

2016-01-01

Leptospirosis is a neglected infectious disease caused by spirochetes from the genus Leptospira . Pathogenic microorganisms, notably those which reach the blood circulation such as Leptospira , have evolved multiple strategies to escape the host complement system, which is important for innate and acquired immunity. Leptospira avoid complement-mediated killing through: (i) recruitment of host complement regulators; (ii) acquisition of host proteases that cleave complement proteins on the bacterial surface; and, (iii) secretion of proteases that inactivate complement proteins in the Leptospira surroundings. The recruitment of host soluble complement regulatory proteins includes the acquisition of Factor H (FH) and FH-like-1 (alternative pathway), C4b-binding protein (C4BP) (classical and lectin pathways), and vitronectin (Vn) (terminal pathway). Once bound to the leptospiral surface, FH and C4BP retain cofactor activity of Factor I in the cleavage of C3b and C4b, respectively. Vn acquisition by leptospires may result in terminal pathway inhibition by blocking C9 polymerization. The second evasion mechanism lies in plasminogen (PLG) binding to the leptospiral surface. In the presence of host activators, PLG is converted to enzymatically active plasmin, which is able to degrade C3b, C4b, and C5 at the surface of the pathogen. A third strategy used by leptospires to escape from complement system is the active secretion of proteases. Pathogenic, but not saprophytic leptospires, are able to secrete metalloproteases that cleave C3 (central complement molecule), Factor B (alternative pathway), and C4 and C2 (classical and lectin pathways). The purpose of this review is to fully explore these complement evasion mechanisms, which act together to favor Leptospira survival and multiplication in the host.

Complement Evasion by Pathogenic Leptospira

PubMed Central

Fraga, Tatiana Rodrigues; Isaac, Lourdes; Barbosa, Angela Silva

2016-01-01

Leptospirosis is a neglected infectious disease caused by spirochetes from the genus Leptospira. Pathogenic microorganisms, notably those which reach the blood circulation such as Leptospira, have evolved multiple strategies to escape the host complement system, which is important for innate and acquired immunity. Leptospira avoid complement-mediated killing through: (i) recruitment of host complement regulators; (ii) acquisition of host proteases that cleave complement proteins on the bacterial surface; and, (iii) secretion of proteases that inactivate complement proteins in the Leptospira surroundings. The recruitment of host soluble complement regulatory proteins includes the acquisition of Factor H (FH) and FH-like-1 (alternative pathway), C4b-binding protein (C4BP) (classical and lectin pathways), and vitronectin (Vn) (terminal pathway). Once bound to the leptospiral surface, FH and C4BP retain cofactor activity of Factor I in the cleavage of C3b and C4b, respectively. Vn acquisition by leptospires may result in terminal pathway inhibition by blocking C9 polymerization. The second evasion mechanism lies in plasminogen (PLG) binding to the leptospiral surface. In the presence of host activators, PLG is converted to enzymatically active plasmin, which is able to degrade C3b, C4b, and C5 at the surface of the pathogen. A third strategy used by leptospires to escape from complement system is the active secretion of proteases. Pathogenic, but not saprophytic leptospires, are able to secrete metalloproteases that cleave C3 (central complement molecule), Factor B (alternative pathway), and C4 and C2 (classical and lectin pathways). The purpose of this review is to fully explore these complement evasion mechanisms, which act together to favor Leptospira survival and multiplication in the host. PMID:28066433

The TFPI-2 derived peptide EDC34 improves outcome of gram-negative sepsis.

PubMed

Papareddy, Praveen; Kalle, Martina; Sørensen, Ole E; Malmsten, Martin; Mörgelin, Matthias; Schmidtchen, Artur

2013-01-01

Sepsis is characterized by a dysregulated host-pathogen response, leading to high cytokine levels, excessive coagulation and failure to eradicate invasive bacteria. Novel therapeutic strategies that address crucial pathogenetic steps during infection are urgently needed. Here, we describe novel bioactive roles and therapeutic anti-infective potential of the peptide EDC34, derived from the C-terminus of tissue factor pathway inhibitor-2 (TFPI-2). This peptide exerted direct bactericidal effects and boosted activation of the classical complement pathway including formation of antimicrobial C3a, but inhibited bacteria-induced activation of the contact system. Correspondingly, in mouse models of severe Escherichia coli and Pseudomonas aeruginosa infection, treatment with EDC34 reduced bacterial levels and lung damage. In combination with the antibiotic ceftazidime, the peptide significantly prolonged survival and reduced mortality in mice. The peptide's boosting effect on bacterial clearance paired with its inhibiting effect on excessive coagulation makes it a promising therapeutic candidate for invasive Gram-negative infections.

The TFPI-2 Derived Peptide EDC34 Improves Outcome of Gram-Negative Sepsis

PubMed Central

Papareddy, Praveen; Kalle, Martina; Sørensen, Ole E.; Malmsten, Martin; Mörgelin, Matthias; Schmidtchen, Artur

2013-01-01

Sepsis is characterized by a dysregulated host-pathogen response, leading to high cytokine levels, excessive coagulation and failure to eradicate invasive bacteria. Novel therapeutic strategies that address crucial pathogenetic steps during infection are urgently needed. Here, we describe novel bioactive roles and therapeutic anti-infective potential of the peptide EDC34, derived from the C-terminus of tissue factor pathway inhibitor-2 (TFPI-2). This peptide exerted direct bactericidal effects and boosted activation of the classical complement pathway including formation of antimicrobial C3a, but inhibited bacteria-induced activation of the contact system. Correspondingly, in mouse models of severe Escherichia coli and Pseudomonas aeruginosa infection, treatment with EDC34 reduced bacterial levels and lung damage. In combination with the antibiotic ceftazidime, the peptide significantly prolonged survival and reduced mortality in mice. The peptide's boosting effect on bacterial clearance paired with its inhibiting effect on excessive coagulation makes it a promising therapeutic candidate for invasive Gram-negative infections. PMID:24339780

A pore-forming protein implements VLR-activated complement cytotoxicity in lamprey.

PubMed

Wu, Fenfang; Feng, Bo; Ren, Yong; Wu, Di; Chen, Yue; Huang, Shengfeng; Chen, Shangwu; Xu, Anlong

2017-01-01

Lamprey is a basal vertebrate with a unique adaptive immune system, which uses variable lymphocyte receptors (VLRs) for antigen recognition. Our previous study has shown that lamprey possessed a distinctive complement pathway activated by VLR. In this study, we identified a natterin family member-lamprey pore-forming protein (LPFP) with a jacalin-like lectin domain and an aerolysin-like pore-forming domain. LPFP had a high affinity with mannan and could form oligomer in the presence of mannan. LPFP could deposit on the surface of target cells, form pore-like complex resembling a wheel with hub and spokes, and mediate powerful cytotoxicity on target cells. These pore-forming proteins along with VLRs and complement molecules were essential for the specific cytotoxicity against exogenous pathogens and tumor cells. This unique cytotoxicity implemented by LPFP might emerge before or in parallel with the IgG-based classical complement lytic pathway completed by polyC9.

Preventing Ototoxic Synergy of Prior Noise Trauma during Aminoglycoside Therapy

DTIC Science & Technology

2017-06-01

with aminoglycoside antibiotics that have broad-spectrum bactericidal activity for treating or preventing life-threatening infections. However...with aminoglycoside antibiotics that have broad- spectrum bactericidal activity for treating or preventing life-threatening infections. However...aminoglycoside uptake. What was accomplished under these goals? 1) Major activities Due to lab relocation, the project was interrupted in 2015, and

The nature of inherent bactericidal activity: insights from the nanotopology of three species of dragonfly

NASA Astrophysics Data System (ADS)

Mainwaring, David E.; Nguyen, Song Ha; Webb, Hayden; Jakubov, Timur; Tobin, Mark; Lamb, Robert N.; Wu, Alex H.-F.; Marchant, Richard; Crawford, Russell J.; Ivanova, Elena P.

2016-03-01

While insect wings are widely recognised as multi-functional, recent work showed that this extends to extensive bactericidal activity brought about by cell deformation and lysis on the wing nanotopology. We now quantitatively show that subtle changes to this topography result in substantial changes in bactericidal activity that are able to span an order of magnitude. Notably, the chemical composition of the lipid nanopillars was seen by XPS and synchrotron FTIR microspectroscopy to be similar across these activity differences. Modelling the interaction between bacterial cells and the wing surface lipids of 3 species of dragonflies, that inhabit similar environments, but with distinctly different behavioural repertoires, provided the relationship between surface structure and antibacterial functionality. In doing so, these principal behavioural patterns correlated with the demands for antimicrobial efficiency dictated by differences in their foraging strategies. This work now reveals a new feature in the design elegance of natural multi-functional surfaces as well providing insights into the bactericidal mechanism underlying inherently antimicrobial materials, while suggesting that nanotopology is related to the evolutionary development of a species through the demands of its behavioural repertoire. The underlying relationship between the processes of wetting, adhesion and capillarity of the lipid nanopillars and bactericidal efficiency suggests new prospects for purely mechano-responsive antibacterial surfaces.While insect wings are widely recognised as multi-functional, recent work showed that this extends to extensive bactericidal activity brought about by cell deformation and lysis on the wing nanotopology. We now quantitatively show that subtle changes to this topography result in substantial changes in bactericidal activity that are able to span an order of magnitude. Notably, the chemical composition of the lipid nanopillars was seen by XPS and synchrotron FTIR microspectroscopy to be similar across these activity differences. Modelling the interaction between bacterial cells and the wing surface lipids of 3 species of dragonflies, that inhabit similar environments, but with distinctly different behavioural repertoires, provided the relationship between surface structure and antibacterial functionality. In doing so, these principal behavioural patterns correlated with the demands for antimicrobial efficiency dictated by differences in their foraging strategies. This work now reveals a new feature in the design elegance of natural multi-functional surfaces as well providing insights into the bactericidal mechanism underlying inherently antimicrobial materials, while suggesting that nanotopology is related to the evolutionary development of a species through the demands of its behavioural repertoire. The underlying relationship between the processes of wetting, adhesion and capillarity of the lipid nanopillars and bactericidal efficiency suggests new prospects for purely mechano-responsive antibacterial surfaces. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr08542j

Antibody to a conserved antigenic target is protective against diverse prokaryotic and eukaryotic pathogens

PubMed Central

Cywes-Bentley, Colette; Skurnik, David; Zaidi, Tanweer; Roux, Damien; DeOliveira, Rosane B.; Garrett, Wendy S.; Lu, Xi; O’Malley, Jennifer; Kinzel, Kathryn; Zaidi, Tauqeer; Rey, Astrid; Perrin, Christophe; Fichorova, Raina N.; Kayatani, Alexander K. K.; Maira-Litràn, Tomas; Gening, Marina L.; Tsvetkov, Yury E.; Nifantiev, Nikolay E.; Bakaletz, Lauren O.; Pelton, Stephen I.; Golenbock, Douglas T.; Pier, Gerald B.

2013-01-01

Microbial capsular antigens are effective vaccines but are chemically and immunologically diverse, resulting in a major barrier to their use against multiple pathogens. A β-(1→6)–linked poly-N-acetyl-d-glucosamine (PNAG) surface capsule is synthesized by four proteins encoded in genetic loci designated intercellular adhesion in Staphylococcus aureus or polyglucosamine in selected Gram-negative bacterial pathogens. We report that many microbial pathogens lacking an identifiable intercellular adhesion or polyglucosamine locus produce PNAG, including Gram-positive, Gram-negative, and fungal pathogens, as well as protozoa, e.g., Trichomonas vaginalis, Plasmodium berghei, and sporozoites and blood-stage forms of Plasmodium falciparum. Natural antibody to PNAG is common in humans and animals and binds primarily to the highly acetylated glycoform of PNAG but is not protective against infection due to lack of deposition of complement opsonins. Polyclonal animal antibody raised to deacetylated glycoforms of PNAG and a fully human IgG1 monoclonal antibody that both bind to native and deacetylated glycoforms of PNAG mediated complement-dependent opsonic or bactericidal killing and protected mice against local and/or systemic infections by Streptococcus pyogenes, Streptococcus pneumoniae, Listeria monocytogenes, Neisseria meningitidis serogroup B, Candida albicans, and P. berghei ANKA, and against colonic pathology in a model of infectious colitis. PNAG is also a capsular polysaccharide for Neisseria gonorrhoeae and nontypable Hemophilus influenzae, and protects cells from environmental stress. Vaccination targeting PNAG could contribute to immunity against serious and diverse prokaryotic and eukaryotic pathogens, and the conserved production of PNAG suggests that it is a critical factor in microbial biology. PMID:23716675

In vivo activation of equine eosinophils and neutrophils by experimental Strongylus vulgaris infections.

PubMed

Dennis, V A; Klei, T R; Chapman, M R; Jeffers, G W

1988-12-01

Eosinophils and neutrophils from ponies with Strongylus vulgaris-induced eosinophilia (eosinophilic ponies; activated eosinophils and neutrophils) were assayed in vitro for chemotactic and chemokinetic responses to zymosan-activated serum (ZAS) using the filter system in Boyden chambers, for Fc and complement (C) receptors using the EA and EAC-rosette assays, respectively, and for phagocytic and bactericidal activities using opsonized Escherichia coli and the acridine orange method. The responses of activated eosinophils and neutrophils in the above assays were compared with those of eosinophils and neutrophils from S. vulgaris-naive ponies without eosinophilia (noneosinophilic ponies; nonactivated eosinophils and neutrophils). Differences in cell density following centrifugation in a continuous Percoll gradient were used to further characterize the heterogeneity of activated eosinophils and neutrophils. Activated and nonactivated eosinophils demonstrated similar chemotactic responses to ZAS while activated and nonactivated neutrophils demonstrated similar chemokinetic responses to ZAS. A higher percentage of activated eosinophils and neutrophils expressed Fc and C receptors compared with nonactivated cells (P less than 0.05). Generally, higher percentages of eosinophils and neutrophils expressed C than Fc receptors. However, the percentage of neutrophils with both receptors was higher than that of eosinophils. Phagocytosis and killing of E. coli by either type of eosinophil were not consistently observed. Both activated and nonactivated neutrophils phagocytized E. coli and significant differences between the two cell types were not observed. The bacterial activity, however, of activated neutrophils was significantly greater than that obtained using nonactivated neutrophils (P less than 0.05). Activated eosinophils and neutrophils were both separated into two distinct fractions based on differences in cell densities. A higher percentage of band 2 eosinophils (density of 1.106) expressed C receptors than did band 1 eosinophils (density of 1.049) (P less than 0.05). A higher percentage of band 1 neutrophils (density of 1.072) expressed both Fc and C receptors and these neutrophils were more phagocytic and bactericidal than were band 2 neutrophils (density of 1.082) (P less than 0.05). These data suggest that equine eosinophils and neutrophils are activated by chronic S. vulgaris infections.

In vitro and bactericidal activities of ABT-492, a novel fluoroquinolone, against Gram-positive and Gram-negative organisms.

PubMed

Almer, Laurel S; Hoffrage, Jennifer B; Keller, Erika L; Flamm, Robert K; Shortridge, Virginia D

2004-07-01

In vitro activities of ABT-492, ciprofloxacin, levofloxacin, trovafloxacin, moxifloxacin, gatifloxacin, and gemifloxacin were compared. ABT-492 was more potent against quinolone-susceptible and -resistant gram-positive organisms, had activity similar to that of ciprofloxacin against certain members of the family Enterobacteriaceae, and had comparable activity against quinolone-susceptible, nonfermentative, gram-negative organisms. Bactericidal activity of ABT-492 was also evaluated.

Complement in Lupus Nephritis: New Perspectives.

PubMed

Bao, Lihua; Cunningham, Patrick N; Quigg, Richard J

2015-09-01

Systemic lupus erythematosus (SLE) is an autoimmune disorder caused by loss of tolerance to self-antigens, the production of autoantibodies and deposition of complement-fixing immune complexes (ICs) in injured tissues. SLE is characterized by a wide range of clinical manifestations and targeted organs, with lupus nephritis being one of the most serious complications. The complement system consists of three pathways and is tightly controlled by a set of regulatory proteins to prevent injudicious complement activation on host tissue. The involvement of the complement system in the pathogenesis of SLE is well accepted; yet, its exact role is still not clear. Complement plays dual roles in the pathogenesis of SLE. On the one hand, the complement system appears to have protective features in that hereditary homozygous deficiencies of classical pathway components, such as C1q and C4, are associated with an increased risk for SLE. On the other hand, IC-mediated activation of complement in affected tissues is clearly evident in both experimental and human SLE along with pathological features that are logical consequences of complement activation. Studies in genetically altered mice have shown that lack of complement inhibitors, such as complement factor H (CFH) or decay-accelerating factor (DAF) accelerates the development of experimental lupus nephritis, while treatment with recombinant protein inhibitors, such as Crry-Ig, CR2-Crry, CR2-DAF and CR2-CFH, ameliorates the disease development. Complement-targeted drugs, including soluble complement receptor 1 (TP10), C1 esterase inhibitor and a monoclonal anti-C5 antibody (eculizumab), have been shown to inhibit complement safely, and are now being investigated in a variety of clinical conditions. SLE is an autoimmune disorder which targets multiple systems. Complement is centrally involved and plays dual roles in the pathogenesis of SLE. Studies from experimental lupus models and clinical trials support the use of complement-targeted therapy in the treatment of SLE.

Evaluation of antimicrobial peptides as novel bactericidal agents for room temperature-stored platelets.

PubMed

Mohan, Ketha V K; Rao, Shilpakala Sainath; Atreya, Chintamani D

2010-01-01

A single cost-effective pathogen inactivation approach would help to improve the safety of our nation's blood supply. Several methods and technologies are currently being studied to help reduce bacterial contamination of blood components. There is clearly need for simple and easy-to-use pathogen inactivation techniques specific to plasma, platelets (PLTs), and red blood cells. In this report, we introduce a novel proof of concept: using known therapeutic antimicrobial peptides (AMPs) as bactericidal agents for room temperature-stored PLT concentrates (PCs). Nine synthetic AMPs, four from PLT microbicidal protein-derived peptides (PD1-4) and five Arg-Trp (RW) repeat peptides containing one to five repeats, were tested for bactericidal activity in plasma and PC samples spiked with Staphylococcus aureus, S. epidermidis, Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, and Bacillus cereus. A 3-log reduction of viable bacteria was considered as the bactericidal activity of a given peptide. In both plasma alone and PCs, RW3 peptide demonstrated bactericidal activity against S. aureus, S. epidermidis, E. coli, P. aeruginosa, and K. pneumoniae; PD4 and RW2 against P. aeruginosa; and RW4 against K. pneumoniae. The activity of each of these four peptides against the remaining bacterial species in the test panel resulted in less than a 3-log reduction in the number of viable bacteria and hence considered ineffective. These findings suggest a new approach to improving the safety of blood components, demonstrating the potential usefulness of screening therapeutic AMPs against selected bacteria to identify suitable bactericidal agents for stored plasma, PCs, and other blood products.

Immunological response to nitroglycerin-loaded shear-responsive liposomes in vitro and in vivo.

PubMed

Buscema, Marzia; Matviykiv, Sofiya; Mészáros, Tamás; Gerganova, Gabriela; Weinberger, Andreas; Mettal, Ute; Mueller, Dennis; Neuhaus, Frederik; Stalder, Etienne; Ishikawa, Takashi; Urbanics, Rudolf; Saxer, Till; Pfohl, Thomas; Szebeni, János; Zumbuehl, Andreas; Müller, Bert

2017-10-28

Liposomes formulated from the 1,3-diamidophospholipid Pad-PC-Pad are shear-responsive and thus promising nano-containers to specifically release a vasodilator at stenotic arteries. The recommended preclinical safety tests for therapeutic liposomes of nanometer size include the in vitro assessment of complement activation and the evaluation of the associated risk of complement activation-related pseudo-allergy (CARPA) in vivo. For this reason, we measured complement activation by Pad-PC-Pad formulations in human and porcine sera, along with the nanopharmaceutical-mediated cardiopulmonary responses in pigs. The evaluated formulations comprised of Pad-PC-Pad liposomes, with and without polyethylene glycol on the surface of the liposomes, and nitroglycerin as a model vasodilator. The nitroglycerin incorporation efficiency ranged from 25% to 50%. In human sera, liposome formulations with 20mg/mL phospholipid gave rise to complement activation, mainly via the alternative pathway, as reflected by the rises in SC5b-9 and Bb protein complex concentrations. Formulations having a factor of ten lower phospholipid content did not result in measurable complement activation. The weak complement activation induced by Pad-PC-Pad liposomal formulations was confirmed by the results obtained by performing an in vivo study in a porcine model, where hemodynamic parameters were monitored continuously. Our study suggests that, compared to FDA-approved liposomal drugs, Pad-PC-Pad exhibits less or similar risks of CARPA. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Alternative pathway regulation by factor H modulates Streptococcus pneumoniae induced proinflammatory cytokine responses by decreasing C5a receptor crosstalk.

PubMed

van der Maten, Erika; de Bont, Cynthia M; de Groot, Ronald; de Jonge, Marien I; Langereis, Jeroen D; van der Flier, Michiel

2016-12-01

Bacterial pathogens not only stimulate innate immune receptors, but also activate the complement system. Crosstalk between complement C5a receptor (C5aR) and other innate immune receptors is known to enhance the proinflammatory cytokine response. An important determinant of the magnitude of complement activation is the activity of the alternative pathway, which serves as an amplification mechanism for complement activation. Both alternative pathway activity as well as plasma levels of factor H, a key inhibitor of the alternative pathway, show large variation within the human population. Here, we studied the effect of factor H-mediated regulation of the alternative pathway on bacterial-induced proinflammatory cytokine responses. We used the human pathogen Streptococcus pneumoniae as a model stimulus to induce proinflammatory cytokine responses in human peripheral blood mononuclear cells. Serum containing active complement enhanced pneumococcal induced proinflammatory cytokine production through C5a release and C5aR crosstalk. We found that inhibition of the alternative pathway by factor H, with a concentration equivalent to a high physiological level, strongly reduced C5a levels and decreased proinflammatory cytokine production in human peripheral blood mononuclear cells. This suggests that variation in alternative pathway activity due to variation in factor H plasma levels affects individual cytokine responses during infection. Copyright © 2016 Elsevier Ltd. All rights reserved.

Bactericidal activity of propylene glycol, glycerine, polyethylene glycol 400, and polyethylene glycol 1000 against selected microorganisms

PubMed Central

Nalawade, Triveni Mohan; Bhat, Kishore; Sogi, Suma H. P.

2015-01-01

Aim: The aim of the present study was to evaluate the bactericidal activity of propylene glycol, glycerine, polyethylene glycol 400 (PEG 400), and polyethylene glycol 1000 (PEG 1000) against selected microorganisms in vitro. Materials and Methods: Five vehicles, namely propylene glycol, glycerine, PEG 400, PEG 1000, and combination of propylene glycol with PEG 400, were tested for their bactericidal activity. The minimum bactericidal concentration was noted against four standard strains of organisms, i.e. Streptococcus mutans American Type Culture Collection (ATCC) 25175, Streptococcus mutans ATCC 12598, Enterococcus faecalis ATCC 35550, and Escherichia coli ATCC 25922, using broth dilution assay. Successful endodontic therapy depends upon thorough disinfection of root canals. In some refractory cases, routine endodontic therapy is not sufficient, so intracanal medicaments are used for proper disinfection of canals. Intracanal medicaments are dispensed with vehicles which aid in increased diffusion through the dentinal tubules and improve their efficacy. Among the various vehicles used, glycerine is easily available, whereas others like propylene glycol and polyethylene glycol have to be procured from appropriate sources. Also, these vehicles, being viscous, aid in sustained release of the medicaments and improve their handling properties. The most commonly used intracanal medicaments like calcium hydroxide are ineffective on many microorganisms, while most of the other medicaments like MTAD (Mixture of Tetracycline, an Acid, and a Detergent) and Triple Antibiotic Paste (TAP) consist of antibiotics which can lead to development of antibiotic resistance among microorganisms. Thus, in order to use safer and equally effective intracanal medicaments, newer alternatives like chlorhexidine gluconate, ozonized water, etc., are being explored. Similarly, the five vehicles mentioned above are being tested for their antimicrobial activity in this study. Results: All vehicles exhibited bactericidal activity at 100% concentration. Conclusion: Propylene glycol was effective against three organisms namely S. mutans E. faecalis and E. coli and its bactericidal activity was at 50%, 25% and 50% respectively. PEG 1000 was effective against S. mutans and E. coli at 25%. Hence propylene glycol was effective on more number of organisms of which E. faecalis is a known resistant species. PEG 1000 was bactericidal at a lower concentration but was effective on two organisms only. PMID:25992336

Bactericidal activity of propylene glycol, glycerine, polyethylene glycol 400, and polyethylene glycol 1000 against selected microorganisms.

PubMed

Nalawade, Triveni Mohan; Bhat, Kishore; Sogi, Suma H P

2015-01-01

The aim of the present study was to evaluate the bactericidal activity of propylene glycol, glycerine, polyethylene glycol 400 (PEG 400), and polyethylene glycol 1000 (PEG 1000) against selected microorganisms in vitro. Five vehicles, namely propylene glycol, glycerine, PEG 400, PEG 1000, and combination of propylene glycol with PEG 400, were tested for their bactericidal activity. The minimum bactericidal concentration was noted against four standard strains of organisms, i.e. Streptococcus mutans American Type Culture Collection (ATCC) 25175, Streptococcus mutans ATCC 12598, Enterococcus faecalis ATCC 35550, and Escherichia coli ATCC 25922, using broth dilution assay. Successful endodontic therapy depends upon thorough disinfection of root canals. In some refractory cases, routine endodontic therapy is not sufficient, so intracanal medicaments are used for proper disinfection of canals. Intracanal medicaments are dispensed with vehicles which aid in increased diffusion through the dentinal tubules and improve their efficacy. Among the various vehicles used, glycerine is easily available, whereas others like propylene glycol and polyethylene glycol have to be procured from appropriate sources. Also, these vehicles, being viscous, aid in sustained release of the medicaments and improve their handling properties. The most commonly used intracanal medicaments like calcium hydroxide are ineffective on many microorganisms, while most of the other medicaments like MTAD (Mixture of Tetracycline, an Acid, and a Detergent) and Triple Antibiotic Paste (TAP) consist of antibiotics which can lead to development of antibiotic resistance among microorganisms. Thus, in order to use safer and equally effective intracanal medicaments, newer alternatives like chlorhexidine gluconate, ozonized water, etc., are being explored. Similarly, the five vehicles mentioned above are being tested for their antimicrobial activity in this study. All vehicles exhibited bactericidal activity at 100% concentration. Propylene glycol was effective against three organisms namely S. mutans E. faecalis and E. coli and its bactericidal activity was at 50%, 25% and 50% respectively. PEG 1000 was effective against S. mutans and E. coli at 25%. Hence propylene glycol was effective on more number of organisms of which E. faecalis is a known resistant species. PEG 1000 was bactericidal at a lower concentration but was effective on two organisms only.

Effect of increased CRM₁₉₇ carrier protein dose on meningococcal C bactericidal antibody response.

PubMed

Lee, Lucia H; Blake, Milan S

2012-04-01

New multivalent CRM(197)-based conjugate vaccines are available for childhood immunization. Clinical studies were reviewed to assess meningococcal group C (MenC) antibody responses following MenC-CRM(197) coadministration with CRM(197)-based pneumococcal or Haemophilus influenzae type b conjugate vaccines. Infants receiving a total CRM(197) carrier protein dose of ∼50 μg and concomitant diphtheria-tetanus-acellular pertussis (DTaP)-containing vaccine tended to have lower MenC geometric mean antibody titers and continued to have low titers after the toddler dose. Nevertheless, at least 95% of children in the reported studies achieved a MenC serum bactericidal antibody (SBA) titer of ≥ 1:8 after the last infant or toddler dose. SBA was measured using an assay with a baby rabbit or human complement source. Additional studies are needed to assess long-term antibody persistence and MenC CRM(197) conjugate vaccine immunogenicity using alternative dosing schedules.

Persistence of the immune response after MenACWY-CRM vaccination and response to a booster dose, in adolescents, children and infants.

PubMed

Baxter, Roger; Keshavan, Pavitra; Welsch, Jo Anne; Han, Linda; Smolenov, Igor

2016-05-03

Persistence of bactericidal antibodies following vaccination is extremely important for protection against invasive meningococcal disease, given the epidemiology and rapid progression of meningococcal infection. We present an analysis of antibody persistence and booster response to MenACWY-CRM, in adolescents, children and infants, from 7 clinical studies. Immunogenicity was assessed using the serum bactericidal assay with both human and rabbit complement. Post-vaccination hSBA titers were high, with an age- and serogroup-specific decline in titers up to 1 y and stable levels up to 5 y The waning of hSBA titers over time was more pronounced among infants and toddlers and the greatest for serogroup A. However, rSBA titers against serogroup A were consistently higher and showed little decline over time, suggesting that protection against this serogroup may be sustained. A single booster dose of MenACWY-CRM administered at 3 to 5 y induced a robust immune response in all age groups.

Binding of Complement Factor H (FH) Decreases Protective Anti-FH Binding Protein Antibody Responses of Infant Rhesus Macaques Immunized With a Meningococcal Serogroup B Vaccine

PubMed Central

Granoff, Dan M.; Costa, Isabella; Konar, Monica; Giuntini, Serena; Van Rompay, Koen K. A.; Beernink, Peter T.

2015-01-01

Background. The meningococcal vaccine antigen, factor H (FH)–binding protein (FHbp), binds human complement FH. In human FH transgenic mice, binding decreased protective antibody responses. Methods. To investigate the effect of primate FH binding, we immunized rhesus macaques with a 4-component serogroup B vaccine (4CMenB). Serum FH in 6 animals bound strongly to FHbp (FHbp-FHhigh) and, in 6 animals, bound weakly to FHbp (FHbp-FHlow). Results. There were no significant differences between the respective serum bactericidal responses of the 2 groups against meningococcal strains susceptible to antibody to the NadA or PorA vaccine antigens. In contrast, anti-FHbp bactericidal titers were 2-fold lower in FHbp-FHhigh macaques against a strain with an exact FHbp match to the vaccine (P = .08) and were ≥4-fold lower against 4 mutants with other FHbp sequence variants (P ≤ .005, compared with FHbp-FHlow macaques). Unexpectedly, postimmunization sera from all 12 macaques enhanced FH binding to meningococci. In contrast, serum anti-FHbp antibodies elicited by 4CMenB in mice whose mouse FH did not bind to the vaccine antigen inhibited FH binding. Conclusions. Binding of FH to FHbp decreases protective anti-FHbp antibody responses of macaques to 4CMenB. Even low levels of FH binding skew the antibody repertoire to FHbp epitopes outside of the FH-binding site, which enhance FH binding. PMID:25676468

Targeting C-reactive protein for the treatment of cardiovascular disease

NASA Astrophysics Data System (ADS)

Pepys, Mark B.; Hirschfield, Gideon M.; Tennent, Glenys A.; Ruth Gallimore, J.; Kahan, Melvyn C.; Bellotti, Vittorio; Hawkins, Philip N.; Myers, Rebecca M.; Smith, Martin D.; Polara, Alessandra; Cobb, Alexander J. A.; Ley, Steven V.; Andrew Aquilina, J.; Robinson, Carol V.; Sharif, Isam; Gray, Gillian A.; Sabin, Caroline A.; Jenvey, Michelle C.; Kolstoe, Simon E.; Thompson, Darren; Wood, Stephen P.

2006-04-01

Complement-mediated inflammation exacerbates the tissue injury of ischaemic necrosis in heart attacks and strokes, the most common causes of death in developed countries. Large infarct size increases immediate morbidity and mortality and, in survivors of the acute event, larger non-functional scars adversely affect long-term prognosis. There is thus an important unmet medical need for new cardioprotective and neuroprotective treatments. We have previously shown that human C-reactive protein (CRP), the classical acute-phase protein that binds to ligands exposed in damaged tissue and then activates complement, increases myocardial and cerebral infarct size in rats subjected to coronary or cerebral artery ligation, respectively. Rat CRP does not activate rat complement, whereas human CRP activates both rat and human complement. Administration of human CRP to rats is thus an excellent model for the actions of endogenous human CRP. Here we report the design, synthesis and efficacy of 1,6-bis(phosphocholine)-hexane as a specific small-molecule inhibitor of CRP. Five molecules of this palindromic compound are bound by two pentameric CRP molecules, crosslinking and occluding the ligand-binding B-face of CRP and blocking its functions. Administration of 1,6-bis(phosphocholine)-hexane to rats undergoing acute myocardial infarction abrogated the increase in infarct size and cardiac dysfunction produced by injection of human CRP. Therapeutic inhibition of CRP is thus a promising new approach to cardioprotection in acute myocardial infarction, and may also provide neuroprotection in stroke. Potential wider applications include other inflammatory, infective and tissue-damaging conditions characterized by increased CRP production, in which binding of CRP to exposed ligands in damaged cells may lead to complement-mediated exacerbation of tissue injury.

Interaction of metronidazole with DNA repair mutants of Escherichia coli.

PubMed

Yeung, T C; Beaulieu, B B; McLafferty, M A; Goldman, P

1984-01-01

It has been proposed that one of metronidazole's partially reduced intermediates interacts either with DNA to exert a bactericidal effect or with water to form acetamide. To test this hypothesis we have examined the effect of metronidazole on several mutants of Escherichia coli that are defective in DNA repair. UV-susceptible RecA- and UvrB- point mutants have an increased susceptibility to metronidazole as manifested by both a decreased minimal inhibitory concentration and a greater bactericidal response to metronidazole in resting cultures. By these criteria, however, we find that UvrB- deletion mutants, which lack the ability to reduce nitrate and chlorate, are no more susceptible to metronidazole than is the wild type. We find, however, that these deletion mutants also lack the ability to reduce metronidazole and thus possibly to form its reactive species. When metronidazole's bactericidal effect is expressed in terms of the concurrent accumulation of acetamide derived from metronidazole, then all RecA- and UvrB- mutants are killed more efficiently than their wild types. The data are consistent, therefore, with metronidazole's lethal effect being mediated by a partially reduced intermediate on the metabolic pathway between metronidazole and acetamide. Defects in other aspects of the DNA repair system do not confer this increased susceptibility to the proposed intermediate. A Tag- mutant, for example, which is defective in 3-methyl-adenine-DNA glycosylase, does not have this increased susceptibility to the presumed precursor of acetamide. Thus, these results provide further support for the hypothesis that the bactericidal effect of metronidazole is mediated by a partially reduced intermediate in the metabolic conversion of metronidazole to acetamide and suggest that this intermediate interacts with DNA to produce a lesion similar to that caused by UV light.

Interaction of metronidazole with DNA repair mutants of Escherichia coli.

PubMed Central

Yeung, T C; Beaulieu, B B; McLafferty, M A; Goldman, P

1984-01-01

It has been proposed that one of metronidazole's partially reduced intermediates interacts either with DNA to exert a bactericidal effect or with water to form acetamide. To test this hypothesis we have examined the effect of metronidazole on several mutants of Escherichia coli that are defective in DNA repair. UV-susceptible RecA- and UvrB- point mutants have an increased susceptibility to metronidazole as manifested by both a decreased minimal inhibitory concentration and a greater bactericidal response to metronidazole in resting cultures. By these criteria, however, we find that UvrB- deletion mutants, which lack the ability to reduce nitrate and chlorate, are no more susceptible to metronidazole than is the wild type. We find, however, that these deletion mutants also lack the ability to reduce metronidazole and thus possibly to form its reactive species. When metronidazole's bactericidal effect is expressed in terms of the concurrent accumulation of acetamide derived from metronidazole, then all RecA- and UvrB- mutants are killed more efficiently than their wild types. The data are consistent, therefore, with metronidazole's lethal effect being mediated by a partially reduced intermediate on the metabolic pathway between metronidazole and acetamide. Defects in other aspects of the DNA repair system do not confer this increased susceptibility to the proposed intermediate. A Tag- mutant, for example, which is defective in 3-methyl-adenine-DNA glycosylase, does not have this increased susceptibility to the presumed precursor of acetamide. Thus, these results provide further support for the hypothesis that the bactericidal effect of metronidazole is mediated by a partially reduced intermediate in the metabolic conversion of metronidazole to acetamide and suggest that this intermediate interacts with DNA to produce a lesion similar to that caused by UV light. PMID:6367636

Membrane attack complex of complement is not essential for immune mediated demyelination in experimental autoimmune neuritis.

PubMed

Tran, Giang T; Hodgkinson, Suzanne J; Carter, Nicole M; Killingsworth, Murray; Nomura, Masaru; Verma, Nirupama D; Plain, Karren M; Boyd, Rochelle; Hall, Bruce M

2010-12-15

Antibody deposition and complement activation, especially membrane attack complex (MAC) formation are considered central for immune mediated demyelination. To examine the role of MAC in immune mediated demyelination, we studied experimental allergic neuritis (EAN) in Lewis rats deficient in complement component 6 (C6) that cannot form MAC. A C6 deficient Lewis (Lewis/C6-) strain of rats was bred by backcrossing the defective C6 gene, from PVG/C6- rats, onto the Lewis background. Lewis/C6- rats had the same C6 gene deletion as PVG/C6- rats and their sera did not support immune mediated haemolysis unless C6 was added. Active EAN was induced in Lewis and Lewis/C6- rats by immunization with bovine peripheral nerve myelin in complete Freund's adjuvant (CFA), and Lewis/C6- rats had delayed clinical EAN compared to the Lewis rats. Peripheral nerve demyelination in Lewis/C6- was also delayed but was similar in extent at the peak of disease. Compared to Lewis, Lewis/C6- nerves had no MAC deposition, reduced macrophage infiltrate and IL-17A, but similar T cell infiltrate and Th1 cytokine mRNA expression. ICAM-1 and P-selectin mRNA expression and immunostaining on vascular endothelium were delayed in Lewis C6- compared to Lewis rats' nerves. This study found that MAC was not required for immune mediated demyelination; but that MAC enhanced early symptoms and early demyelination in EAN, either by direct lysis or by sub-lytic induction of vascular endothelial expression of ICAM-1 and P-selectin. Copyright © 2010 Elsevier B.V. All rights reserved.

Sophora flavescens protects against mycobacterial Trehalose Dimycolate-induced lung granuloma by inhibiting inflammation and infiltration of macrophages.

PubMed

Liu, Dehua; Chan, Ben Chung-Lap; Cheng, Ling; Tsang, Miranda Sin-Man; Zhu, Jing; Wong, Chun-Wai; Jiao, Delong; Chan, Helen Yau-Tsz; Leung, Ping Chung; Lam, Christopher Wai-Kei; Wong, Chun Kwok

2018-03-02

The immune system responds to Mycobacterium tuberculosis (MTB) infection by forming granulomas to quarantine the bacteria from spreading. Granuloma-mediated inflammation is a cause of lung destruction and disease transmission. Sophora flavescens (SF) has been demonstrated to exhibit bactericidal activities against MTB. However, its immune modulatory activities on MTB-mediated granulomatous inflammation have not been reported. In the present study, we found that flavonoids from Sophora flavescens (FSF) significantly suppressed the pro-inflammatory mediators released from mouse lung alveolar macrophages (MH-S) upon stimulation by trehalose dimycolate (TDM), the most abundant lipoglycan on MTB surface. Moreover, FSF reduced adhesion molecule (LFA-1) expression on MH-S cells after TDM stimulation. Furthermore, FSF treatment on TDM-activated lung epithelial (MLE-12) cells significantly downregulated macrophage chemoattractant protein (MCP-1/CCL2) expression, which in turn reduced the in vitro migration of MH-S to MLE-12 cells. In addition, FSF increased the clearance of mycobacterium bacteria (Mycobacterium aurum) in macrophages. FSF mainly affected the Mincle-Syk-Erk signaling pathway in TDM-activated MH-S cells. In TDM-induced mouse granulomas model, oral administration with FSF significantly suppressed lung granulomas formation and inflammation. These findings collectively implicated an anti-inflammatory role of FSF on MTB-mediated granulomatous inflammation, thereby providing evidence of FSF as an efficacious adjunct treatment during mycobacterial infection.

Clinical roundtable monograph: Paroxysmal nocturnal hemoglobinuria: a case-based discussion.

PubMed

Szer, Jeff; Hill, Anita; Weitz, Ilene Ceil

2012-11-01

Paroxysmal nocturnal hemoglobinuria (PNH) is a rare, acquired disorder characterized by chronic intravascular hemolysis as the primary clinical manifestation and morbidities that include anemia, thrombosis, renal impairment, pulmonary hypertension, and bone marrow failure. The prevalence of the PNH clone (from <1-100% PNH granulocytes) is approximately 16 per million, and careful monitoring is required. The average age of onset of the clinical disease is the early 30s, although it can present at all ages. PNH is caused by the acquisition of a somatic mutation of the gene phosphatidylinositol glycan anchor (PIG-A) in a multipotent hematopoietic stem cell (HSC), with clonal expansion of the mutated HSC. The mutation causes a deficiency in the synthesis of glycosylphosphatidylinositol (GPI). In cells derived from normal HSCs, the complement regulatory proteins CD55 and CD59 are anchored to the hematopoietic cell membrane surface via GPI, protecting the cells from complement-mediated lysis. However, in patients with PNH, these 2 proteins, along with numerous other GPI-linked proteins, are absent from the cell surface of red cells, granulocytes, monocytes, and platelets, resulting in complement-mediated intravascular hemolysis and other complications. Lysis of red blood cells is the most obvious manifestation, but as other cell lineages are also affected, this complement-mediated attack contributes to additional complications, such as thrombosis. Eculizumab, a humanized monoclonal antibody against the C5 complement protein, is the only effective drug therapy for PNH patients. The antibody prevents cleavage of the C5 protein by C5 convertase, in turn preventing generation of C5b-9 and release of C5a, thereby protecting from hemolysis of cells lacking the CD59 surface protein and other complications associated with complement activation. Drs. Ilene C. Weitz, Anita Hill, and Jeff Szer discuss 3 recent cases of patients with PNH.

Inactivation of virus in solution by cold atmospheric pressure plasma: identification of chemical inactivation pathways

NASA Astrophysics Data System (ADS)

Aboubakr, Hamada A.; Gangal, Urvashi; Youssef, Mohammed M.; Goyal, Sagar M.; Bruggeman, Peter J.

2016-05-01

Cold atmospheric pressure plasma (CAP) inactivates bacteria and virus through in situ production of reactive oxygen and nitrogen species (RONS). While the bactericidal and virucidal efficiency of plasmas is well established, there is limited knowledge about the chemistry leading to the pathogen inactivation. This article describes a chemical analysis of the CAP reactive chemistry involved in the inactivation of feline calicivirus. We used a remote radio frequency CAP produced in varying gas mixtures leading to different plasma-induced chemistries. A study of the effects of selected scavengers complemented with positive control measurements of relevant RONS reveal two distinctive pathways based on singlet oxygen and peroxynitrous acid. The first mechanism is favored in the presence of oxygen and the second in the presence of air when a significant pH reduction is induced in the solution by the plasma. Additionally, smaller effects of the H2O2, O3 and \\text{NO}2- produced were also found. Identification of singlet oxygen-mediated 2-imidazolone/2-oxo-His (His  +14 Da)—an oxidative modification of His 262 comprising the capsid protein of feline calicivirus links the plasma induced singlet oxygen chemistry to viral inactivation.

Bactericidal Activity of Usnic Acid-Loaded Electrospun Fibers.

PubMed

Araújo, Evando S; Pereira, Eugênia C; da Costa, Mateus M; da Silva, Nicácio H; de Oliveira, Helinando P

2016-01-01

Usnic acid has been progressively reported in the literature as one of the most important lichen metabolites characterized by a rich diversity of applications such as antifungal, antimicrobial, antiprotozoal and antiviral agent. Particularly, antimicrobial activity of usnic acid can be improved by encapsulation of active molecules in enteric electrospun fibers, allowing the controlled release of active molecule at specific pH. Few relevant patents to the topic have been reviewed and cited. Bactericidal activity of usnic acid-loaded electrospun fibers of Eudragit L-100 and polyvinylpyrrolidone was examined against Staphylococcus aureus using inhibition hales methodology. The controlled release of active material at high pH is established after 10 minutes of interaction with media and results in reasonable activity against S. aureus, as detected by inhibition hales. The strong biological activity of usnic acid-loaded electrospun fibers provides a promising application for corresponding material as a bactericidal agent for wound healing treatment.

Pleiotropic Effects of Cell Wall Amidase LytA on Streptococcus pneumoniae Sensitivity to the Host Immune Response

PubMed Central

Ramos-Sevillano, Elisa; Urzainqui, Ana; Campuzano, Susana; Moscoso, Miriam; González-Camacho, Fernando; Domenech, Mirian; Rodríguez de Córdoba, Santiago; Sánchez-Madrid, Francisco; Brown, Jeremy S.; García, Ernesto

2014-01-01

The complement system is a key component of the host immune response for the recognition and clearance of Streptococcus pneumoniae. In this study, we demonstrate that the amidase LytA, the main pneumococcal autolysin, inhibits complement-mediated immunity independently of effects on pneumolysin by a complex process of impaired complement activation, increased binding of complement regulators, and direct degradation of complement C3. The use of human sera depleted of either C1q or factor B confirmed that LytA prevented activation of both the classical and alternative pathways, whereas pneumolysin inhibited only the classical pathway. LytA prevented binding of C1q and the acute-phase protein C-reactive protein to S. pneumoniae, thereby reducing activation of the classical pathway on the bacterial surface. In addition, LytA increased recruitment of the complement downregulators C4BP and factor H to the pneumococcal cell wall and directly cleaved C3b and iC3b to generate degradation products. As a consequence, C3b deposition and phagocytosis increased in the absence of LytA and were markedly enhanced for the lytA ply double mutant, confirming that a combination of LytA and Ply is essential for the establishment of pneumococcal pneumonia and sepsis in a murine model of infection. These data demonstrate that LytA has pleiotropic effects on complement activation, a finding which, in combination with the effects of pneumolysin on complement to assist with pneumococcal complement evasion, confirms a major role of both proteins for the full virulence of the microorganism during septicemia. PMID:25404032

In Vitro and Bactericidal Activities of ABT-492, a Novel Fluoroquinolone, against Gram-Positive and Gram-Negative Organisms

PubMed Central

Almer, Laurel S.; Hoffrage, Jennifer B.; Keller, Erika L.; Flamm, Robert K.; Shortridge, Virginia D.

2004-01-01

In vitro activities of ABT-492, ciprofloxacin, levofloxacin, trovafloxacin, moxifloxacin, gatifloxacin, and gemifloxacin were compared. ABT-492 was more potent against quinolone-susceptible and -resistant gram-positive organisms, had activity similar to that of ciprofloxacin against certain members of the family Enterobacteriaceae, and had comparable activity against quinolone-susceptible, nonfermentative, gram-negative organisms. Bactericidal activity of ABT-492 was also evaluated. PMID:15215148

Translational Mini-Review Series on Complement Factor H: Structural and functional correlations for factor H

PubMed Central

Schmidt, C Q; Herbert, A P; Hocking, H G; Uhrín, D; Barlow, P N

2008-01-01

The 155-kDa glycoprotein, complement factor H (CFH), is a regulator of complement activation that is abundant in human plasma. Three-dimensional structures of over half the 20 complement control protein (CCP) modules in CFH have been solved in the context of single-, double- and triple-module segments. Proven binding sites for C3b occupy the N and C termini of this elongated molecule and may be brought together by a bend in CFH mediated by its central CCP modules. The C-terminal CCP 20 is key to the ability of the molecule to adhere to polyanionic markers on self-surfaces where CFH acts to regulate amplification of the alternative pathway of complement. The surface patch on CCP 20 that binds to model glycosaminoglycans has been mapped using nuclear magnetic resonance (NMR), as has a second glycosaminoglycan-binding patch on CCP 7. These patches include many of the residue positions at which sequence variations have been linked to three complement-mediated disorders: dense deposit disease, age-related macular degeneration and atypical haemolytic uraemic syndrome. In one plausible model, CCP 20 anchors CFH to self-surfaces via a C3b/polyanion composite binding site, CCP 7 acts as a ‘proof-reader’ to help discriminate self- from non-self patterns of sulphation, and CCPs 1–4 disrupt C3/C5 convertase formation and stability. PMID:18081691

On the first polyarsenic organic compound from nature: arsenicin A from the New Caledonian marine sponge Echinochalina bargibanti.

PubMed

Mancini, Ines; Guella, Graziano; Frostin, Maryvonne; Hnawia, Edouard; Laurent, Dominique; Debitus, Cecile; Pietra, Francesco

2006-12-04

Reported here is the first polyarsenic compound ever found in nature. Denominated arsenicin A, it was isolated along a bioassay-guided fractionation of the organic extract of the poecilosclerid sponge Echinochalina bargibanti collected from the north-eastern coast of New Caledonia. In defining an adamantine-type polyarsenic structure for this compound, deceptively simple NMR spectra were complemented by extensive mass spectral analysis. However, it was only the synthesis of a model compound that provided the basis to discriminate structure 4 from other spectrally compatible structures for arsenicin A; to this end, a comparative ab initio simulation of IR spectra for the natural and the synthetic compounds was decisive. Arsenicin A is endowed with potent bactericidal and fungicidal activities on human pathogenic strains. All this may revive pharmacological interest in arsenic compounds while prompting us to rethink the arsenic cycle in nature.

High-Mobility Group Box 1-Induced Complement Activation Causes Sterile Inflammation.

PubMed

Kim, Sook Young; Son, Myoungsun; Lee, Sang Eun; Park, In Ho; Kwak, Man Sup; Han, Myeonggil; Lee, Hyun Sook; Kim, Eun Sook; Kim, Jae-Young; Lee, Jong Eun; Choi, Ji Eun; Diamond, Betty; Shin, Jeon-Soo

2018-01-01

High-mobility group box 1 (HMGB1), a well-known danger-associated molecular pattern molecule, acts as a pro-inflammatory molecule when secreted by activated immune cells or released after necrotic cell damage. HMGB1 binds to immunogenic bacterial components and augments septic inflammation. In this study, we show how HMGB1 mediates complement activation, promoting sterile inflammation. We show that HMGB1 activates the classical pathway of complement system in an antibody-independent manner after binding to C1q. The C3a complement activation product in human plasma and C5b-9 membrane attack complexes on cell membrane surface are detected after the addition of HMGB1. In an acetaminophen (APAP)-induced hepatotoxicity model, APAP injection reduced HMGB1 levels and elevated C3 levels in C1q-deficient mouse serum samples, compared to that in wild-type (WT) mice. APAP-induced C3 consumption was inhibited by sRAGE treatment in WT mice. Moreover, in a mouse model of brain ischemia-reperfusion injury based on middle cerebral arterial occlusion, C5b-9 complexes were deposited on vessels where HMGB1 was accumulated, an effect that was suppressed upon HMGB1 neutralization. We propose that the HMGB1 released after cell necrosis and in ischemic condition can trigger the classical pathway of complement activation to exacerbate sterile inflammation.

High-Mobility Group Box 1-Induced Complement Activation Causes Sterile Inflammation

PubMed Central

Kim, Sook Young; Son, Myoungsun; Lee, Sang Eun; Park, In Ho; Kwak, Man Sup; Han, Myeonggil; Lee, Hyun Sook; Kim, Eun Sook; Kim, Jae-Young; Lee, Jong Eun; Choi, Ji Eun; Diamond, Betty; Shin, Jeon-Soo

2018-01-01

High-mobility group box 1 (HMGB1), a well-known danger-associated molecular pattern molecule, acts as a pro-inflammatory molecule when secreted by activated immune cells or released after necrotic cell damage. HMGB1 binds to immunogenic bacterial components and augments septic inflammation. In this study, we show how HMGB1 mediates complement activation, promoting sterile inflammation. We show that HMGB1 activates the classical pathway of complement system in an antibody-independent manner after binding to C1q. The C3a complement activation product in human plasma and C5b-9 membrane attack complexes on cell membrane surface are detected after the addition of HMGB1. In an acetaminophen (APAP)-induced hepatotoxicity model, APAP injection reduced HMGB1 levels and elevated C3 levels in C1q-deficient mouse serum samples, compared to that in wild-type (WT) mice. APAP-induced C3 consumption was inhibited by sRAGE treatment in WT mice. Moreover, in a mouse model of brain ischemia–reperfusion injury based on middle cerebral arterial occlusion, C5b-9 complexes were deposited on vessels where HMGB1 was accumulated, an effect that was suppressed upon HMGB1 neutralization. We propose that the HMGB1 released after cell necrosis and in ischemic condition can trigger the classical pathway of complement activation to exacerbate sterile inflammation. PMID:29696019

Characterization of a Complement-Binding Protein, DRS, from Strains of Streptococcus pyogenes Containing the emm12 and emm55 Genes

PubMed Central

Binks, Michael; Sriprakash, K. S.

2004-01-01

An extracellular protein of Streptococcus pyogenes, streptococcal inhibitor of complement (SIC), and its variant, called DRS (distantly related to SIC), are expressed by some S. pyogenes strains. SIC from type 1 (M1) isolates of S. pyogenes interferes with complement-mediated cell lysis, reportedly via its interaction with complement proteins. In this study we demonstrate that S. pyogenes strains carrying emm12 and emm55 (the genes for the M12 and M55 proteins, respectively) express and secrete DRS. This protein, like SIC, binds to the C6 and C7 complement proteins, and competition enzyme-linked immunosorbent assay experiments demonstrate that DRS competes with SIC for C6 and C7 binding. Similarly, SIC competes with DRS for binding to the complement proteins. Despite this, the recombinant DRS preparation showed no significant effect on complement function, as determined by lysis of sensitized sheep erythrocytes. Furthermore, the presence of DRS is not inhibitory to SIC activity. PMID:15213143

Characterization of a complement-binding protein, DRS, from strains of Streptococcus pyogenes containing the emm12 and emm55 genes.

PubMed

Binks, Michael; Sriprakash, K S

2004-07-01

An extracellular protein of Streptococcus pyogenes, streptococcal inhibitor of complement (SIC), and its variant, called DRS (distantly related to SIC), are expressed by some S. pyogenes strains. SIC from type 1 (M1) isolates of S. pyogenes interferes with complement-mediated cell lysis, reportedly via its interaction with complement proteins. In this study we demonstrate that S. pyogenes strains carrying emm12 and emm55 (the genes for the M12 and M55 proteins, respectively) express and secrete DRS. This protein, like SIC, binds to the C6 and C7 complement proteins, and competition enzyme-linked immunosorbent assay experiments demonstrate that DRS competes with SIC for C6 and C7 binding. Similarly, SIC competes with DRS for binding to the complement proteins. Despite this, the recombinant DRS preparation showed no significant effect on complement function, as determined by lysis of sensitized sheep erythrocytes. Furthermore, the presence of DRS is not inhibitory to SIC activity.

Effectiveness of disinfectants used in cooling towers against Legionella pneumophila.

PubMed

García, M T; Pelaz, C

2008-01-01

Legionella persists in man-made aquatic installations despite preventive treatments. More information about disinfectants could improve the effectiveness of treatments. This study tests the susceptibility of Legionella pneumophila serogroup (sg) 1 against 8 disinfectants used in cooling tower treatments. We determined the minimal inhibitory concentration (MIC), minimal bactericidal concentration (MBC) and bactericidal effect of sodium hypochlorite (A), hydrogen peroxide with silver nitrate (B), didecyldimethylammonium chloride (C), benzalkonium chloride (D), tributyltetradecylphosphonium chloride (E), tetrahydroxymethylphosphonium sulfide (F), 2,2-dibromonitropropionamide (G) and chloromethylisothiazolone (H) against 28 L. pneumophila sg 1 isolates. MIC and MBC values were equivalent. Bacteria are less susceptible to disinfectants F, B, D and A than to H, E, C and G. All disinfectants induced a bactericidal effect. The effect rate is dose dependent for G, H, F and B; the effect is fast for the rest of disinfectants at any concentration. The bactericidal activity of disinfectants A, G and F depends on the susceptibility test used. All disinfectants have bactericidal activity against L. pneumophila sg 1 at concentrations used in cooling tower treatments. Results depend on the assay for some products.

Glucose-6-Phosphate Dehydrogenase Deficiency Mimicking Atypical Hemolytic Uremic Syndrome.

PubMed

Walsh, Patrick R; Johnson, Sally; Brocklebank, Vicky; Salvatore, Jacobo; Christian, Martin; Kavanagh, David

2018-02-01

A 4-year-old boy presented with nonimmune hemolysis, thrombocytopenia, and acute kidney injury. Investigations for an underlying cause failed to identify a definitive cause and a putative diagnosis of complement-mediated atypical hemolytic uremic syndrome (aHUS) was made. The patient was started initially on plasma exchange and subsequently eculizumab therapy, after which his kidney function rapidly improved. While on eculizumab therapy, despite adequate complement blockade, he presented 2 more times with hemolytic anemia and thrombocytopenia, but without renal involvement. Genetic analysis did not uncover a mutation in any known aHUS gene (CFH, CFI, CFB, C3, CD46, THBD, INF2, and DGKE) and anti-factor H antibodies were undetectable. Whole-exome sequencing was undertaken to identify a cause for the eculizumab resistance. This revealed a pathogenic variant in G6PD (glucose-6-phosphate dehydrogenase), which was confirmed by functional analysis demonstrating decreased erythrocyte G6PD activity. Eculizumab therapy was withdrawn. Complement-mediated aHUS is a diagnosis of exclusion and this case highlights the diagnostic difficulty that remains without an immediately available biomarker for confirmation. This case of G6PD deficiency presented with a phenotype clinically indistinguishable from complement-mediated aHUS. We recommend that G6PD deficiency be included in the differential diagnosis of patients presenting with aHUS and suggest measuring erythrocyte G6PD concentrations in these patients. Copyright © 2017. Published by Elsevier Inc.

Complement Activation on Platelets Correlates with a Decrease in Circulating Immature Platelets in Patients with Immune Thrombocytopenic Purpura

PubMed Central

Peerschke, Ellinor I.B.; Andemariam, Biree; Yin, Wei; Bussel, James B.

2010-01-01

The role of the complement system in immune thrombocytopenic purpura (ITP) is not well defined. We examined plasma from 79 patients with ITP, 50 healthy volunteers, and 25 patients with non-immune mediated thrombocytopenia, to investigate their complement activation/fixation capacity (CAC) on immobilized heterologous platelets. Enhanced CAC was found in 46 plasma samples (59%) from patients with ITP, but no samples from patients with non-immune mediated thrombocytopenia. Plasma from healthy volunteers was used for comparison. In patients with ITP, an enhanced plasma CAC was associated with a decreased circulating absolute immature platelet fraction (A-IPF) (<15 × 109/L) (p = 0.027) and thrombocytopenia (platelet count less than 100K/μl) (p= 0.024). The positive predictive value of an enhanced CAC for a low A-IPF was 93%, with a specificity of 77%. The specificity and positive predictive values increased to 100% when plasma CAC was defined strictly by enhanced C1q and/or C4d deposition on test platelets. Although no statistically significant correlation emerged between CAC and response to different pharmacologic therapies, an enhanced response to splenectomy was noted (p <0.063). Thus, complement fixation may contribute to the thrombocytopenia of ITP by enhancing clearance of opsonized platelets from the circulation, and/or directly damaging platelets and megakaryocytes. PMID:19925495

Antibody-Mediated Rejection in Human Cardiac Allografts: Evaluation of Immunoglobulins and Complement Activation Products C4d and C3d as Markers

PubMed Central

Rodriguez, E. R.; Skojec, Diane V.; Tan, Carmela D.; Zachary, Andrea A.; Kasper, Edward K.; Conte, John V.; Baldwin, William M.

2005-01-01

Antibody-mediated rejection (AMR) in human heart transplantation is an immunopathologic process in which injury to the graft is in part the result of activation of complement and it is poorly responsive to conventional therapy. We evaluated by immunofluorescence (IF), 665 consecutive endomyocardial biopsies from 165 patients for deposits of immunoglobulins and complement. Diffuse IF deposits in a linear capillary pattern greater than 2+ were considered significant. Clinical evidence of graft dysfunction was correlated with complement deposits. IF 2+ or higher was positive for IgG, 66%; IgM, 12%; IgA, 0.6%; C1q, 1.8%; C4d, 9% and C3d, 10%. In 3% of patients, concomitant C4d and C3d correlated with graft dysfunction or heart failure. In these 5 patients AMR occurred 56–163 months after transplantation, and they responded well to therapy for AMR but not to treatment with steroids. Systematic evaluation of endomyocardial biopsies is not improved by the use of antibodies for immunoglobulins or C1q. Concomitant use of C4d and C3d is very useful to diagnose AMR, when correlated with clinical parameters of graft function. AMR in heart transplant patients can occur many months or years after transplant. PMID:16212640

C1 inhibitor-mediated myocardial protection from chronic intermittent hypoxia-induced injury

PubMed Central

Fu, Jinrong; Guo, Furong; Chen, Cheng; Yu, Xiaoman; Hu, Ke; Li, Mingjiang

2016-01-01

The optimal treatment for chronic intermittent hypoxia (CIH)-induced cardiovascular injuries has yet to be determined. The aim of the current study was to explore the potential protective effect and mechanism of a C1 inhibitor in CIH in the myocardium. The present study used a rat model of CIH in which complement regulatory protein, known as C1 inhibitor (C1INH), was administered to the rats in the intervention groups. Cardiomyocyte apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling. The expression of proteins associated with the apoptotic pathway, such as B-cell lymphoma 2 (Bcl-2), Bax and caspase-3 were detected by western blot analysis. The expression of complement C3 protein and RNA were also analyzed. C1INH was observed to improve the cardiac function in rats with CIH. Myocardial myeloperoxidase activity, a marker of neutrophil infiltration, was significantly decreased in the C1INH intervention group compared with the CIH control group, and cardiomyocyte apoptosis was significantly attenuated (P<0.05). Western blotting and reverse transcription-polymerase chain reaction analysis indicated that the protein expression levels of Bcl-2 were decreased and those of Bax were increased in the CIH group compared with the normal control group, but the protein expression levels of Bcl-2 were increased and those of Bax were decreased in the C1INH intervention group, as compared with the CIH group. Furthermore, the CIH-induced expression and synthesis of complement C3 in the myocardium were also reduced in the C1INH intervention group. C1INH, in addition to inhibiting complement activation and inflammation, preserved cardiac function in CIH-mediated myocardial cell injury through an anti-apoptotic mechanism. PMID:27698713

Bothrops asper snake venom and its metalloproteinase BaP-1 activate the complement system. Role in leucocyte recruitment.

PubMed Central

Farsky, S H; Gonçalves, L R; Gutiérrez, J M; Correa, A P; Rucavado, A; Gasque, P; Tambourgi, D V

2000-01-01

The venom of the snake Bothrops asper, the most important poisonous snake in Central America, evokes an inflammatory response, the mechanisms of which are not well characterized. The objectives of this study were to investigate whether B. asper venom and its purified toxins--phospholipases and metalloproteinase--activate the complement system and the contribution of the effect on leucocyte recruitment. In vitro chemotaxis assays were performed using Boyden's chamber model to investigate the ability of serum incubated with venom and its purified toxins to induce neutrophil migration. The complement consumption by the venom was evaluated using an in vitro haemolytic assay. The importance of complement activation by the venom on neutrophil migration was investigated in vivo by injecting the venom into the peritoneal cavity of C5-deficient mice. Data obtained demonstrated that serum incubated with crude venom and its purified metalloproteinase BaP-1 are able to induce rat neutrophil chemotaxis, probably mediated by agent(s) derived from the complement system. This hypothesis was corroborated by the capacity of the venom to activate this system in vitro. The involvement of C5a in neutrophil chemotaxis induced by venom-activated serum was demonstrated by abolishing migration when neutrophils were pre-incubated with antirat C5a receptor antibody. The relevance of the complement system in in vivo leucocyte mobilization was further demonstrated by the drastic decrease of this response in C5-deficient mice. Pre-incubation of serum with the soluble human recombinant complement receptor type 1 (sCR 1) did not prevent the response induced by the venom, but abolished the migration evoked by metalloproteinase-activated serum. These data show the role of the complement system in bothropic envenomation and the participation of metalloproteinase in the effect. Also, they suggest that the venom may contain other component(s) which can cause direct activation of C5a. PMID:11200361

A whole blood bactericidal assay for tuberculosis.

PubMed

Wallis, R S; Palaci, M; Vinhas, S; Hise, A G; Ribeiro, F C; Landen, K; Cheon, S H; Song, H Y; Phillips, M; Dietze, R; Ellner, J J

2001-04-15

The bactericidal activity of orally administered antituberculosis (anti-TB) drugs was determined in a whole blood culture model of intracellular infection in which microbial killing reflects the combined effects of drug and immune mechanisms. Rifampin (Rif) was the most active compound studied and reduced the number of viable bacilli by >4 logs. Isoniazid (INH), 2 quinolones, and pyrazinamide (PZA) showed intermediate levels of activity. Ethambutol exerted only a bacteristatic effect; amoxicillin/clavulanate was inactive. The combination of INH-Rif-PZA showed strong activity against 11 drug-sensitive isolates (mean, -3.8 log) but no activity against 12 multidrug-resistant (MDR) strains. The combination of levofloxacin-PZA-ethambutol had intermediate bactericidal activity against MDR isolates (mean, -1.2 log) but failed to equal that of INH-Rif-PZA against sensitive isolates (P<.001). The whole blood BACTEC method (Becton Dickinson) may be useful for the early clinical evaluation of new anti-TB drugs and in the management of individual patients.

Ibrutinib interferes with the cell-mediated anti-tumor activities of therapeutic CD20 antibodies: implications for combination therapy

PubMed Central

Roit, Fabio Da; Engelberts, Patrick J.; Taylor, Ronald P.; Breij, Esther C.W.; Gritti, Giuseppe; Rambaldi, Alessandro; Introna, Martino; Parren, Paul W.H.I.; Beurskens, Frank J.; Golay, Josée

2015-01-01

The novel Bruton tyrosine kinase inhibitor ibrutinib and phosphatidyl-4-5-biphosphate 3-kinase-δ inhibitor idelalisib are promising drugs for the treatment of chronic lymphocytic leukemia and B-cell non-Hodgkin lymphoma, either alone or in combination with anti-CD20 antibodies. We investigated the possible positive or negative impact of these drugs on all known mechanisms of action of both type I and type II anti-CD20 antibodies. Pretreatment with ibrutinib for 1 hour did not increase direct cell death of cell lines or chronic lymphocytic leukemia samples mediated by anti-CD20 antibodies. Pre-treatment with ibrutinib did not inhibit complement activation or complement-mediated lysis. In contrast, ibrutinib strongly inhibited all cell-mediated mechanisms induced by anti-CD20 antibodies rituximab, ofatumumab or obinutuzumab, either in purified systems or whole blood assays. Activation of natural killer cells, and antibody-dependent cellular cytotoxicity by these cells, as well as phagocytosis by macrophages or neutrophils were inhibited by ibrutinib with a half maximal effective concentration of 0.3–3 μM. Analysis of anti-CD20 mediated activation of natural killer cells isolated from patients on continued oral ibrutinib treatment suggested that repeated drug dosing inhibits these cells in vivo. Finally we show that the phosphatidyl-4-5-biphosphate 3-kinase-δ inhibitor idelalisib similarly inhibited the immune cell-mediated mechanisms induced by anti-CD20 antibodies, although the effects of this drug at 10 μM were weaker than those observed with ibrutinib at the same concentration. We conclude that the design of combined treatment schedules of anti-CD20 antibodies with these kinase inhibitors should consider the multiple negative interactions between these two classes of drugs. PMID:25344523

Deposition of mannose-binding lectin and ficolins and activation of the lectin pathway of complement on the surface of polyurethane tubing used for cardiopulmonary bypass.

PubMed

Eppa, Łukasz; Pągowska-Klimek, Izabela; Świerzko, Anna S; Moll, Maciej; Krajewski, Wojciech R; Cedzyński, Maciej

2018-04-01

The artificial surface used for cardiopulmonary bypass (CPB) is a crucial factor activating the complement system and thus contributing to the generation of a systemic inflammatory response. The activation of classical and alternative pathways on this artificial surface is well known. In contrast, lectin pathway (LP) activation has not been fully investigated, although noted during CPB in several studies. Moreover, we have recently proved the contribution of the LP to the generation of the systemic inflammatory response syndrome after pediatric cardiac surgery. The aim of this study was to assess LP-mediated complement activation on the surface of polyurethane CPB circuit tubing (noncoated Chalice ® ), used for CPB procedures in children with congenital heart disease. We found deposition of mannose-binding lectin, ficolin-1, -2, and -3 on the surface of unused tubing and on tubing used for CPB from a small minority of patients. Furthermore, we observed deposition of complement C4 activation products on tubing used for CPB and previously unused tubing after incubation with normal serum. The latter finding indicates LP activation in vitro on the polyurethane surface. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 1202-1208, 2018. © 2017 Wiley Periodicals, Inc.

Identification of a central role for complement in osteoarthritis

PubMed Central

Wang, Qian; Rozelle, Andrew L.; Lepus, Christin M.; Scanzello, Carla R.; Song, Jason J.; Larsen, D. Meegan; Crish, James F.; Bebek, Gurkan; Ritter, Susan Y.; Lindstrom, Tamsin M.; Hwang, Inyong; Wong, Heidi H.; Punzi, Leonardo; Encarnacion, Angelo; Shamloo, Mehrdad; Goodman, Stuart B.; Wyss-Coray, Tony; Goldring, Steven R.; Banda, Nirmal K.; Thurman, Joshua M.; Gobezie, Reuben; Crow, Mary K.; Holers, V. Michael; Lee, David M.; Robinson, William H.

2011-01-01

Osteoarthritis, characterized by the breakdown of articular cartilage in synovial joints, has long been viewed as the result of “wear and tear”1. Although low-grade inflammation is detected in osteoarthritis, its role is unclear2–4. Here we identify a central role for the inflammatory complement system in the pathogenesis of osteoarthritis. Through proteomic and transcriptomic analyses of synovial fluids and membranes from individuals with osteoarthritis, we find that expression and activation of complement is abnormally high in human osteoarthritic joints. Using mice genetically deficient in C5, C6, or CD59a, we show that complement, and specifically the membrane attack complex (MAC)-mediated arm of complement, is critical to the development of arthritis in three different mouse models of osteoarthritis. Pharmacological modulation of complement in wild-type mice confirmed the results obtained with genetically deficient mice. Expression of inflammatory and degradative molecules was lower in chondrocytes from destabilized joints of C5-deficient mice than C5-sufficient mice, and MAC induced production of these molecules in cultured chondrocytes. Furthermore, MAC co-localized with matrix metalloprotease (MMP)-13 and with activated extracellular signal-regulated kinase (ERK) around chondrocytes in human osteoarthritic cartilage. Our findings indicate that dysregulation of complement in synovial joints plays a critical role in the pathogenesis of osteoarthritis. PMID:22057346

Green-Tea and Epigallocatechin-3-Gallate are Bactericidal against Bacillus anthracis

DTIC Science & Technology

2017-06-13

EGCG, catechins such 245 as epigallocatechin and epicatechin gallate are also antibacterial agents. Moreover, the 246 bactericidal activity of green...Sharma A, Gupta S, Sarethy IP, Dang S, Gabrani R. 2012. Green tea extract: possible mechanism 285 and antibacterial activity on skin pathogens. Food...was shown to be responsible for this activity , against 30 both the attenuated B. anthracis ANR and the virulent, encapsulated strain B. anthracis

Therapeutic inhibition of the complement system. Y2K update.

PubMed

Asghar, S S; Pasch, M C

2000-09-01

Activation of complement is an essential part of the mechanism of pathogenesis of a large number of human diseases; its inhibition by pharmacological means is likely to suppress disease processes in complement mediated diseases. From this point of view low molecular weight synthetic inhibitors of complement are being developed and high molecular weight natural inhibitors of human origin present in plasma or embedded in cell membrane are being purified or produced in their recombinant forms. This review is concerned with high molecular weight inhibitors, some of which are already in clinical use but may be efficacious in many other diseases in which they have not yet been tried. C1-esterase inhibitor (C1-INH) concentrate prepared from human plasma is being successfully used for the treatment of hereditary angioneurotic edema. Recently, C1-INH has been found to be consumed in severe inflammation and has been shown to exert beneficial effects in several inflammatory conditions such as human sepsis, post-operative myocardial dysfunction due to reperfusion injury, severe capillary leakage syndrome after bone marrow transplantation, reperfusion injury after lung transplantation, burn, and cytotoxicity caused by IL-2 therapy in cancer. Factor I has been used for the treatment of factor I deficiency. Recombinant soluble forms of membrane cofactor protein (MCP), and decay accelerating factor (DAF) have not yet been tried in humans but have been shown to be effective in immune complex mediate inflammation in animals. Organs of pigs transgenic for one or more of human membrane regulators of complement namely membrane cofactor protein (MCP), decay accelerating factor (DAF) or CD59, are being produced for transplantation into humans. They have been shown to be resistant to hyperacute rejection in non-human primates; acute vascular rejection is still a problem in their clinical use. It is hoped that these observations together with future developments will make xeno-transplantation in clinical practice a reality. Several recombinant variants of complement receptor 1 (CR1) have been produced. The most effective of these appears to be sCR1-SLe x, sCR1 part of which inhibits complement and carbohydrate Sle x moiety inhibits selectin mediated interactions of neutrophils and lymphocytes with endothelium. Although clinical trials of sCR1 in humans is eagerly awaited, several of the recombinant versions of sCR1 have been shown to suppress ischemia/reperfusion injury, thermal trauma, and immune complex mediated inflammation. They have also been shown to be effective in experimental models of systemic sclerosis, arthritis, myasthenia gravis, Guillain Barré syndrome and glomerulonephritis. Intravenous immunoglobulin, three of the most prominent properties of which are neutralization of autoantibody activity, suppression of autoantibody production and inhibition of complement activity, is being used in several diseases. These include autoimmune thrombocyopenic purpura, Kawasaki disease and several neurological diseases such as myasthenia gravis and Guillain Barre syndrome. In many uncontrolled small scale studies intravenous immunoglobulin has been shown to be effective in many immunological including dermatological diseases; controlled clinical trials in a large number of patients with these diseases is needed to establish the efficacy. It is hoped that in future therapeutic inhibition of complement will be one of the major approaches to combat many human diseases.

Structure-activity analysis and biological studies of chensinin-1b analogues.

PubMed

Dong, Weibing; Dong, Zhe; Mao, Xiaoman; Sun, Yue; Li, Fei; Shang, Dejing

2016-06-01

Chensinin-1b shows a potent and broad-spectrum bactericidal activity and no hemolytic activity and thus is a potential therapeutic agent against bacterial infection. The NMR structure of chensinin-1b consists of a partially α-helical region (residues 8-14) in a membrane-mimic environment that is distinct from other common antimicrobial peptides. However, further analysis of the structural features of chensinin-1b is required to better understand its bactericidal activity. In this study, a series of N- and C-terminally truncated or amino acid-substituted chensinin-1b analogues were synthesized. Next, the bactericidal activity and bacterial membrane effects of the analogues were investigated. The results indicated that the N-terminal residues play a more significant role than the C-terminal residues in the antimicrobial activity of chensinin-1b. The removal of five amino acids from the C-terminus of chensinin-1b did not affect its biological properties, but helix disruption significantly decreased bactericidal activity. The substitution of positively charged residues increased the helicity and antimicrobial activity of the peptide. We also identified a novel analogue [R(4),R(10)]C1b(3-13) that exhibited similar bactericidal properties with its parent peptide chensinin-1b. Electrostatic interactions between the selected analogues and lipopolysaccharides or cells were detected using isothermal titration calorimetry or zeta potential. The thermodynamic parameters ΔH and ΔS for [R(4),R(10)]C1b(3-13) were -20.48kcalmol(-1) and -0.0408kcalmol(-1)deg(-1), respectively. Chensinin-1b yielded similar results of -26.36kcalmol(-1) and -0.0559kcalmol(-1)deg(-1) for ΔH and ΔS, respectively. These results are consistence with their antimicrobial activities. Lastly, membrane depolarization studies showed that selected analogues exerted bactericidal activity by damaging the cytoplasmic membrane. Antimicrobial peptide chensinin-1b is a candidate for the development of new drugs and a template for the design of synthetic analogues. It mainly exhibits a random coil conformation in membrane environment, and in this manuscript, we characterized the structure of chensinin-1b using NMR spectroscopy, its structure is different than the structures of magainin 2, which has an α-helical conformation and indolicidin, which has a random coil structure. The structural features of chensinin-1b that are required for its potent bactericidal activity were also elucidated. Based on these data, we can fully understand the structure-activity relationship of such peptide and identified a novel analogue with properties that make it an attractive topic for future therapeutic research. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Bactericidal activity and post-antibiotic effect of ozenoxacin against Propionibacterium acnes.

PubMed

Kanayama, Shoji; Okamoto, Kazuaki; Ikeda, Fumiaki; Ishii, Ritsuko; Matsumoto, Tatsumi; Hayashi, Naoki; Gotoh, Naomasa

2017-06-01

Ozenoxacin, a novel non-fluorinated topical quinolone, is used for the treatment of acne vulgaris in Japan. We investigated bactericidal activity and post-antibiotic effect (PAE) of ozenoxacin against Propionibacterium acnes, a major causative bacterium of acne vulgaris. The minimum inhibitory concentrations (MICs) of ozenoxacin against 3 levofloxacin-susceptible strains (MIC of levofloxacin; ≤4 μg/mL) and 3 levofloxacin-resistant strains (MIC of levofloxacin; ≥8 μg/mL) ranged from 0.03 to 0.06 μg/mL and from 0.25 to 0.5 μg/mL, respectively. These MICs of ozenoxacin were almost the same or lower than nadifloxacin and clindamycin. The minimum bactericidal concentrations (MBCs) of ozenoxacin against the levofloxacin-susceptible and -resistant strains were from 0.06 to 8 μg/mL and from 0.5 to 4 μg/mL, respectively. These MBCs were lower than those of nadifloxacin and clindamycin. In time-kill assay, ozenoxacin at 1/4, 1 and 4 times the respective MIC against both levofloxacin-susceptible and -resistant strains showed a concentration-dependent bactericidal activity. Ozenoxacin at 4 times the MICs against the levofloxacin-susceptible strains showed more potent and more rapid onset of bactericidal activity compared to nadifloxacin and clindamycin at 4 times the respective MICs. The PAEs of ozenoxacin at 4 times the MICs against the levofloxacin-susceptible strains were from 3.3 to 17.1 h, which were almost the same or longer than nadifloxacin and clindamycin. In contrast, the PAEs were hardly induced by any antimicrobial agents against the levofloxacin-resistant strains. The present findings suggest that ozenoxacin has a potent bactericidal activity against both levofloxacin-susceptible and -resistant P. acnes, and a long-lasting PAE against levofloxacin-susceptible P. acnes. Copyright © 2017 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

BACTERICIDAL COATINGS ON TEXTILES FOR REMEDIATION OF INTERMICROBE ACTIVITY (BaCTeRIA) SUMMARY REPORT

DTIC Science & Technology

2017-07-07

Activity by Bacillus sp. P11” Food Bioprocess Technol. 4:822- 828. (2011) Levy, S.B and Marshal,l B.; “ Antibacterial resistance worldwide: causes...TEXTILES FOR REMEDIATION OF INTERMICROBE ACTIVITY (BaCTeRIA) SUMMARY REPORT by Tobyn A. Branck Courtney M. Cowell Jennifer M. Rego and...October 2011 – September 2015 4. TITLE AND SUBTITLE BACTERICIDAL COATINGS ON TEXTILES FOR REMEDIATION OF INTERMICROBE ACTIVITY (BaCTeRIA) SUMMARY REPORT

Bactericidal and anti-adhesive properties of culinary and medicinal plants against Helicobacter pylori.

PubMed

O'Mahony, Rachel; Al-Khtheeri, Huda; Weerasekera, Deepaka; Fernando, Neluka; Vaira, Dino; Holton, John; Basset, Christelle

2005-12-21

To investigate the bactericidal and anti-adhesive properties of 25 plants against Helicobacter pylori (H. pylori). Twenty-five plants were boiled in water to produce aqueous extracts that simulate the effect of cooking. The bactericidal activity of the extracts was assessed by a standard kill-curve with seven strains of H. pylori. The anti-adhesive property was assessed by the inhibition of binding of four strains of FITC-labeled H. pylori to stomach sections. Of all the plants tested, eight plants, including Bengal quince, nightshade, garlic, dill, black pepper, coriander, fenugreek and black tea, were found to have no bactericidal effect on any of the isolates. Columbo weed, long pepper, parsley, tarragon, nutmeg, yellow-berried nightshade, threadstem carpetweed, sage and cinnamon had bactericidal activities against H. pylori, but total inhibition of growth was not achieved in this study. Among the plants that killed H. pylori, turmeric was the most efficient, followed by cumin, ginger, chilli, borage, black caraway, oregano and liquorice. Moreover, extracts of turmeric, borage and parsley were able to inhibit the adhesion of H. pylori strains to the stomach sections. Several plants that were tested in our study had bactericidal and/or anti-adhesive effects on H. pylori. Ingestion of the plants with anti-adhesive properties could therefore provide a potent alternative therapy for H. pylori infection, which overcomes the problem of resistance associated with current antibiotic treatment.

Cytotoxic activity against human neuroblastoma and melanoma cells mediated by IgM antibodies derived from peripheral blood of healthy donors.

PubMed

Devarapu, Satish Kumar; Mamidi, Srinivas; Plöger, Frank; Dill, Othmar; Blixt, Ola; Kirschfink, Michael; Schwartz-Albiez, Reinhard

2016-06-15

A small percentage of healthy donors identified in the Western population carry antibodies in their peripheral blood which convey cytotoxic activity against certain human melanoma and neuroblastoma cell lines. We measured the cytotoxic activity of sera and plasmas from healthy donors on the human neuroblastoma cell line Kelly and various melanoma cell lines. Antibodies of IgM isotype, presumably belonging to the class of naturally occurring antibodies, exerted cytotoxic activity in a complement-dependent fashion. Apart from complement-dependent tumor cell lysis, we observed C3 opsonization in all tumor cell lines upon treatment with cytotoxic plasmas. Cell lines tested primarily expressed membrane complement regulatory proteins (mCRP) CD46, CD55 and CD59 to various extents. Blocking of mCRPs by monoclonal antibodies enhanced cell lysis and opsonization, though some melanoma cells remained resistant to complement attack. Epitopes recognized by cytotoxic antibodies were represented by gangliosides such as GD2 and GD3, as evidenced by cellular sialidase pretreatment and enhanced expression of distinct gangliosides. It remains to be clarified why only a small fraction of healthy persons carry these antitumor cytotoxic antibodies. © 2016 UICC.

Effect of amino acid substitution in the staphylococcal peptides warnericin RK and PSMα on their anti-Legionella and hemolytic activities.

PubMed

Marchand, Adrienne; Augenstreich, Jacques; Loiseau, Clémence; Verdon, Julien; Lecomte, Sophie; Berjeaud, Jean-Marc

2015-07-01

Warnericin RK from Staphylococcus warneri and PSMα from Staphylococcus epidermidis are anti-Legionella peptides which were differently classified in a previous study according to their mode of action. Indeed, warnericin RK is highly hemolytic with a bactericidal mode of action, whereas PSMα is poorly hemolytic with a bacteriostatic mode of action toward L. pneumophila. In order to find anti-Legionella peptides which are not hemolytic, a collection of peptides varying in sequence from warnericin RK to PSMα were designed and synthesized, and their anti-Legionella activities, in terms of growth inhibition, permeabilization, and bactericidal effect, as well as their hemolytic activities, were measured and compared. The results showed that some residues, at position 14 for both peptides for instance, were of major importance for bactericidal and hemolytic activities.

[Influence of corynebacteria metabolites on antagonistic activity of H2O2 producing lactobacilli].

PubMed

Bukharin, O V; Sgibnev, A V

2012-01-01

Study combined influence of Corynebacterium genus bacteria metabolites and H2O2 producing lactobacilli on survival rate of Staphylococcus aureus, Escherichia coli and Lactobacillus acidophilus. The ability to inhibit catalase of the test strains used and to reduce bactericidal effect of hydroxyl radical were determined in corynebacteria. H2O2 containing metabolites were obtained by cultivating lactobacilli in mineral medium, the amount of H2O2 was determined by oxidation of TMB by peroxidase. Bactericidal effect of lactobacilli metabolites for test strains treated by corynebacteria metabolites was evaluated by seeding results. Results. Inhibitio by corynebacteria metabolites of S. aureus catalase activity by 30-40% and E. coli catalase activ ity by 40-70% was shown. A reduction of bactericidal effect of hydroxyl radicals by corynebacteria metabolites by 30-35% for S. aureus, 38-42% for E. coli and 70-73% for L. acidophilus was noted. The enchantment of bactericidal effect of lactobacilli after treatment of the test strain by corynebacteria metabolites against S. aureus and E. coli manifested by reduction of the numbe of viable cells by 2-3 lg CFU. For L. acidophilus the bactericidal effect oflactobacilli metabolite in the same conditions reduced, and that led to the increase ofviability by 2-4 lg PFU. A conclusion on the possibility of regulation by associative bacteria the manifestations of antagonistic activity of H2O2 producing dominant microorganisms is made based on the data obtained.

Acquisition of C1 inhibitor by Bordetella pertussis virulence associated gene 8 results in C2 and C4 consumption away from the bacterial surface

PubMed Central

Hovingh, Elise S.; Kuipers, Betsy; Pinelli, Elena; Rooijakkers, Suzan H. M.

2017-01-01

Whooping cough, or pertussis, is a contagious disease of the respiratory tract that is re-emerging worldwide despite high vaccination coverage. The causative agent of this disease is the Gram-negative Bordetella pertussis. Knowledge on complement evasion strategies of this pathogen is limited. However, this is of great importance for future vaccine development as it has become apparent that a novel pertussis vaccine is needed. Here, we unravel the effect of Virulence associated gene 8 (Vag8) of B. pertussis on the human complement system at the molecular level. We show that both recombinant and endogenously secreted Vag8 inhibit complement deposition on the bacterial surface at the level of C4b. We reveal that Vag8 binding to human C1-inhibitor (C1-inh) interferes with the binding of C1-inh to C1s, C1r and MASP-2, resulting in the release of active proteases that subsequently cleave C2 and C4 away from the bacterial surface. We demonstrate that the depletion of these complement components in the bacterial surrounding and subsequent decreased deposition on B. pertussis leads to less complement-mediated bacterial killing. Vag8 is the first protein described that specifically prevents C1s, C1r and MASP-2 binding to C1-inh and thereby mediates complement consumption away from the bacterial surface. Unravelling the mechanism of this unique complement evasion strategy of B. pertussis is one of the first steps towards understanding the interactions between the first line of defense complement and B. pertussis. PMID:28742139

Acquisition of C1 inhibitor by Bordetella pertussis virulence associated gene 8 results in C2 and C4 consumption away from the bacterial surface.

PubMed

Hovingh, Elise S; van den Broek, Bryan; Kuipers, Betsy; Pinelli, Elena; Rooijakkers, Suzan H M; Jongerius, Ilse

2017-07-01

Whooping cough, or pertussis, is a contagious disease of the respiratory tract that is re-emerging worldwide despite high vaccination coverage. The causative agent of this disease is the Gram-negative Bordetella pertussis. Knowledge on complement evasion strategies of this pathogen is limited. However, this is of great importance for future vaccine development as it has become apparent that a novel pertussis vaccine is needed. Here, we unravel the effect of Virulence associated gene 8 (Vag8) of B. pertussis on the human complement system at the molecular level. We show that both recombinant and endogenously secreted Vag8 inhibit complement deposition on the bacterial surface at the level of C4b. We reveal that Vag8 binding to human C1-inhibitor (C1-inh) interferes with the binding of C1-inh to C1s, C1r and MASP-2, resulting in the release of active proteases that subsequently cleave C2 and C4 away from the bacterial surface. We demonstrate that the depletion of these complement components in the bacterial surrounding and subsequent decreased deposition on B. pertussis leads to less complement-mediated bacterial killing. Vag8 is the first protein described that specifically prevents C1s, C1r and MASP-2 binding to C1-inh and thereby mediates complement consumption away from the bacterial surface. Unravelling the mechanism of this unique complement evasion strategy of B. pertussis is one of the first steps towards understanding the interactions between the first line of defense complement and B. pertussis.

Interaction of non-human primate complement and antibodies with hypermucoviscous Klebsiella pneumoniae.

PubMed

Soto, Esteban; Marchi, Sylvia; Beierschmitt, Amy; Kearney, Michael; Francis, Stewart; VanNess, Kimberly; Vandenplas, Michel; Thrall, MaryAnna; Palmour, Roberta

2016-03-08

Emergent hypermucoviscosity (HMV) phenotypes of Klebsiella pneumoniae have been associated with increased invasiveness and pathogenicity in primates. In this study, we investigated the interaction of African green monkeys (AGM) (Chlorocebus aethiops sabaeus) complement and antibody with HMV and non-HMV isolates as in vitro models of primate infection. Significantly greater survival of HMV isolates was evident after incubation in normal serum or whole blood (p < 0.05) of AGM donors when compared to non-HMV strains. Greater survival of HMV strains (p < 0.05) was found after incubation in whole blood and serum from seropositive donors when compared to seronegative donor samples. Additionally, significantly greater amounts of K. pneumoniae were phagocytozed by AGM leukocytes when complement was active (p < 0.05), but no difference in uptake was observed when serum from seropositive or seronegative animals was used in challenged cells utilizing flow cytometry. Results demonstrate that interaction of cellular and humoral immune elements play a role in the in vitro killing of K. pneumoniae, particularly HMV isolates. Neither AGM serum, nor washed whole blood effectively killed HMV isolates; however, assays using heparinized whole blood of seronegative donors significantly reduced viability of HMV and non-HMV strains. The lack of bacterial killing observed in seropositive donors treatments could be at least partially associated with low IgG2 present in these animals. A better understanding of the pathogenesis of klebsiellosis in primates and host immune response is necessary to identify surface molecules that can induce both opsonizing and bactericidal antibody facilitating killing of Klebsiella, and the development of vaccines in human and animals.

Cicada Wing Surface Topography: An Investigation into the Bactericidal Properties of Nanostructural Features.

PubMed

Kelleher, S M; Habimana, O; Lawler, J; O' Reilly, B; Daniels, S; Casey, E; Cowley, A

2016-06-22

Recently, the surface of the wings of the Psaltoda claripennis cicada species has been shown to possess bactericidal properties and it has been suggested that the nanostructure present on the wings was responsible for the bacterial death. We have studied the surface-based nanostructure and bactericidal activity of the wings of three different cicadas (Megapomponia intermedia, Ayuthia spectabile and Cryptotympana aguila) in order to correlate the relationship between the observed surface topographical features and their bactericidal properties. Atomic force microscopy and scanning electron microscopy performed in this study revealed that the tested wing species contained a highly uniform, nanopillar structure on the surface. The bactericidal properties of the cicada wings were investigated by assessing the viability of autofluorescent Pseudomonas fluorescens cells following static adhesion assays and targeted dead/live fluorescence staining through direct microscopic counting methods. These experiments revealed a 20-25% bacterial surface coverage on all tested wing species; however, significant bactericidal properties were observed in the M. intermedia and C. aguila species as revealed by the high dead:live cell ratio on their surfaces. The combined results suggest a strong correlation between the bactericidal properties of the wings and the scale of the nanotopography present on the different wing surfaces.

Complement is a central mediator of radiotherapy-induced tumor-specific immunity and clinical response.

PubMed

Surace, Laura; Lysenko, Veronika; Fontana, Andrea Orlando; Cecconi, Virginia; Janssen, Hans; Bicvic, Antonela; Okoniewski, Michal; Pruschy, Martin; Dummer, Reinhard; Neefjes, Jacques; Knuth, Alexander; Gupta, Anurag; van den Broek, Maries

2015-04-21

Radiotherapy induces DNA damage and cell death, but recent data suggest that concomitant immune stimulation is an integral part of the therapeutic action of ionizing radiation. It is poorly understood how radiotherapy supports tumor-specific immunity. Here we report that radiotherapy induced tumor cell death and transiently activated complement both in murine and human tumors. The local production of pro-inflammatory anaphylatoxins C3a and C5a was crucial to the tumor response to radiotherapy and concomitant stimulation of tumor-specific immunity. Dexamethasone, a drug frequently given during radiotherapy, limited complement activation and the anti-tumor effects of the immune system. Overall, our findings indicate that anaphylatoxins are key players in radiotherapy-induced tumor-specific immunity and the ensuing clinical responses. Copyright © 2015 Elsevier Inc. All rights reserved.

Contribution of the autolysin AtlA to the bactericidal activity of amoxicillin against Enterococcus faecalis JH2-2.

PubMed

Bravetti, Anne-Lise; Mesnage, Stéphane; Lefort, Agnès; Chau, Françoise; Eckert, Catherine; Garry, Louis; Arthur, Michel; Fantin, Bruno

2009-04-01

The bactericidal activity of amoxicillin was investigated against Enterococcus faecalis JH2-2 and against an isogenic mutant deficient in the production of the N-acetylglucosaminidase AtlA. Comparison of the two strains indicated that this autolysin contributes to killing by amoxicillin both in vitro and in a rabbit model of experimental endocarditis.

Structural effect of quaternary ammonium chitin derivatives on their bactericidal activity and specificity.

PubMed

Morkaew, Tirut; Pinyakong, Onruthai; Tachaboonyakiat, Wanpen

2017-08-01

The effect of the quaternary ammonium chitin structure on the bactericidal activity and specificity against Escherichia coli and Staphylococcus aureus was investigated. Quaternary ammonium chitins were synthesized by the separate acylation of chitin (CT) with carboxymethyl trimethylammonium chloride (CMA), 3-carboxypropyl trimethylammonium chloride (CPA) and N-dodecyl-N,N-(dimethylammonio)butyrate (DDMAB). The successful acylation was confirmed by newly formed ester linkage. All three derivatives had a higher surface charge than chitin due to the additional positively charged quaternary ammonium groups. The N-short alkyl substituent (methyl) of CTCMA and CTCPA increased the hydrophilicity whilst the N-long alkyl substituent (dodecyl) of CTDDMAB increased the hydrophobicity compared to chitin. Chitin did not exhibit any bactericidal activity, while CTCMA and CTCPA completely killed E. coli and S. aureus in 30 and 60min, respectively, and CTDDMAB completely killed S. aureus in 10min but did not kill E. coli after a 2-h exposure. Therefore, the N-short alkyl substituent was more effective for killing E. coli and the N-long alkyl substituent conferred specific bactericidal activity against S. aureus. Copyright © 2017 Elsevier B.V. All rights reserved.

The antigenic complex in HIT binds to B cells via complement and complement receptor 2 (CD21)

PubMed Central

Khandelwal, Sanjay; Lee, Grace M.; Hester, C. Garren; Poncz, Mortimer; McKenzie, Steven E.; Sachais, Bruce S.; Rauova, Lubica; Kelsoe, Garnett; Cines, Douglas B.; Frank, Michael

2016-01-01

Heparin-induced thrombocytopenia is a prothrombotic disorder caused by antibodies to platelet factor 4 (PF4)/heparin complexes. The mechanism that incites such prevalent anti-PF4/heparin antibody production in more than 50% of patients exposed to heparin in some clinical settings is poorly understood. To investigate early events associated with antigen exposure, we first examined the interaction of PF4/heparin complexes with cells circulating in whole blood. In healthy donors, PF4/heparin complexes bind preferentially to B cells (>90% of B cells bind to PF4/heparin in vitro) relative to neutrophils, monocytes, or T cells. Binding of PF4 to B cells is heparin dependent, and PF4/heparin complexes are found on circulating B cells from some, but not all, patients receiving heparin. Given the high proportion of B cells that bind PF4/heparin, we investigated complement as a mechanism for noncognate antigen recognition. Complement is activated by PF4/heparin complexes, co-localizes with antigen on B cells from healthy donors, and is present on antigen-positive B cells in patients receiving heparin. Binding of PF4/heparin complexes to B cells is mediated through the interaction between complement and complement receptor 2 (CR2 [CD21]). To the best of our knowledge, these are the first studies to demonstrate complement activation by PF4/heparin complexes, opsonization of PF4/heparin to B cells via CD21, and the presence of complement activation fragments on circulating B cells in some patients receiving heparin. Given the critical contribution of complement to humoral immunity, our observations provide new mechanistic insights into the immunogenicity of PF4/heparin complexes. PMID:27412887

Nonspecific bactericidal activity of the lactoperoxidases-thiocyanate-hydrogen peroxide system of milk against Escherichia coli and some gram-negative pathogens.

PubMed Central

Reiter, B; Marshall, V M; BjörckL; Rosén, C G

1976-01-01

Two strains of Escherichia coli and one strain each of Salmonella typhimurium and Pseudomonas aeruginosa were killed by the bactericidal activity of the lactoperoxidase-thiocyanate-hydrogen peroxide system in milk and in a synthetic medium. H2O2 was supplied exogenously by glucose oxidase, and glucose was produced at a level which was itself noninhibitory. Two phases were distinguished: the first phase was dependent on the oxidation of SCN(-) by lactoperoxidase and H2O2, which was reversed by reducing agent, and the second phase was dependent on the presence of accumulated H2O2, which was reversed by catalase. The latter enzyme could also reverse the first phase, but only when present in excessive and unphysiological levels. The bactericidal activity was greatest at pH 5 and below, and it depended on the SCN(-)concentration and on the number of organisms. Since raw or heated milk neutralizes the acid barrier against infection in the stomach, the bactericidal system discussed may contribute to the prevention of enteric infections in neonates. PMID:5374

Determination of serum bactericidal activity against Escherichia coli by an automated photometric method.

PubMed Central

Crokaert, F; Lismont, M J; van der Linden, M P; Yourassowsky, E

1988-01-01

The resistance of gram-negative bacteria to complement-mediated serum activity is supposedly an important virulence factor. However, the lack of standardization in the methods used to determine serum activity and the many definitions applied make the comparisons between studies very difficult. We developed a rapid photometric method that we compared with a classical killing one. Escherichia coli in the exponential phase of growth in brain heart infusion broth (final inoculum, 10(7) CFU/ml) at 35 degrees C was added to 50% human serum in Veronal buffer. Viable counts and automatic recording of the variations in the optical densities were obtained for 40 E. coli strains isolated from the stools of healthy adults. With the viable count method, 17 (42.5%) were susceptible (at least a 1 log CFU/ml decrease), 17 (42.5%) were resistant (a 0.6 log CFU/ml increase), 4 (10%) were intermediate (poorly growing inoculum or a decrease of less than 1 log CFU/ml), and 2 could not be classified (nonreproducible results). Agreement between both methods was observed for 87.5% of the stool strains. Eight reference strains of known susceptibilities were classified identically by both methods, leading to a final concordance rate of 89.6%. A total of 129 blood isolates were tested by the photometric method: 64 (49.6%) were resistant, 50 (38.8%) were susceptible 5 (3.9%) showed early regrowth, and 10 (7.7%) were not perfectly reproducible. Of these 129 blood isolates, 5 were also tested by the killing method: 37 (49%) were resistant, 32 (43%) were susceptible, and 6 (8%) were intermediate. The concordance rate between both assays was 89% for the blood isolates; when the minor discordances were ruled out, it was 97%. This automated method could be a useful screening tool for detecting resistance to serum in clinical trials and for studying the in vitro variations of this property. PMID:3053761

Targeting complement-mediated immunoregulation for cancer immunotherapy.

PubMed

Kolev, Martin; Markiewski, Maciej M

2018-06-01

Complement was initially discovered as an assembly of plasma proteins "complementing" the cytolytic activity of antibodies. However, our current knowledge places this complex system of several plasma proteins, receptors, and regulators in the center of innate immunity as a bridge between the initial innate responses and adaptive immune reactions. Consequently, complement appears to be pivotal for elimination of pathogens, not only as an early response defense, but by directing the subsequent adaptive immune response. The discovery of functional intracellular complement and its roles in cellular metabolism opened novel avenues for research and potential therapeutic implications. The recent studies demonstrating immunoregulatory functions of complement in the tumor microenvironment and the premetastatic niche shifted the paradigm on our understanding of functions of the complement system in regulating immunity. Several complement proteins, through their interaction with cells in the tumor microenvironment and in metastasis-targeted organs, contribute to modulating tumor growth, antitumor immunity, angiogenesis, and therefore, the overall progression of malignancy and, perhaps, responsiveness of cancer to different therapies. Here, we focus on recent progress in our understanding of immunostimulatory vs. immunoregulatory functions of complement and potential applications of these findings to the design of novel therapies for cancer patients. Copyright © 2018 Elsevier Ltd. All rights reserved.

The effect of polyhexamethylene guanidine hydrochloride (PHMG) derivatives introduced into polylactide (PLA) on the activity of bacterial enzymes.

PubMed

Walczak, Maciej; Richert, Agnieszka; Burkowska-But, Aleksandra

2014-11-01

The present study was aimed at investigating bactericidal properties of polylactide (PLA) films containing three different polyhexamethylene guanidine hydrochloride (PHMG) derivatives and effect of the derivatives on extracellular hydrolytic enzymes and intracellular dehydrogenases. All PHMG derivatives had a slightly stronger bactericidal effect on Staphylococcus aureus than on E. coli but only PHMG granular polyethylene wax (at the concentration of at least 0.6 %) has a bactericidal effect. PHMG derivatives introduced into PLA affected the activity of microbial hydrolases to a small extent. This means that the introduction of PHMG derivatives into PLA will not reduce its enzymatic biodegradation significantly. On the other hand, PHMG derivatives introduced into PLA strongly affected dehydrogenases activity in S. aureus than in E. coli.

PHAGOCYTIN: A BACTERICIDAL SUBSTANCE FROM POLYMORPHONUCLEAR LEUCOCYTES

PubMed Central

Hirsch, James G.

1956-01-01

A technique has been developed for collecting large numbers of polymorphonuclear leucocytes from peritoneal exudates in rabbits. These cells are obtained essentially free from other cell types and from debris. When microphages so procured are disrupted by physical methods and extracted with aqueous salt solutions, the soluble fraction manifests striking bactericidal activity, especially on Gram-negative enteric bacilli. The susceptible microorganisms are not lysed. This bactericidal substance, which has been called phagocytin, appears to be limited in distribution mainly to the polymorphonuclear leucocyte. No phagocytin is present in extracts of rabbit heart, kidney, or skeletal muscle, and rabbit liver and spleen contain much less than do packed leucocytes. Extracts of human and of guinea pig microphages show less bactericidal activity than rabbit cell preparations. Similar extracts of rat and mouse polymorphonuclear leucocytes contain no demonstrable phagocytin. As indicated by its behavior on dialysis, on exposure to proteolytic enzymes, and on salt fractionation, phagocytin appears to be a protein with general properties characteristic of a globulin. It is clearly different from lysozyme and from properdin. Although phagocytin is reasonably stable at temperatures of 65°C. and lower for several hours, solutions of it gradually lose bactericidal activity on standing for prolonged periods at 4°C. This instability, and also the ease with which phagocytin is inactivated, presumably by adsorption, on exposure to a variety of materials, have thus far rendered fruitless efforts to isolate it. PMID:13319580

Bactericidal active ingredient in cryopreserved plasma-treated water with the reduced-pH method for plasma disinfection

NASA Astrophysics Data System (ADS)

Kitano, Katsuhisa; Ikawa, Satoshi; Nakashima, Yoichi; Tani, Atsushi; Yokoyama, Takashi; Ohshima, Tomoko

2016-09-01

For the plasma disinfection of human body, plasma sterilization in liquid is crucial. We found that the plasma-treated water (PTW) has strong bactericidal activity under low pH condition. Physicochemical properties of PTW is discussed based on chemical kinetics. Lower temperature brings longer half-life and the bactericidal activity of PTW can be kept by cryopreservation. High performance PTW, corresponding to the disinfection power of 22 log reduction (B. subtilis spore), can be obtained by special plasma system equipped with cooling device. This is equivalent to 65% H2O2, 14% sodium hypochlorite and 0.33% peracetic acid, which are deadly poison for human. But, it is deactivated soon at higher temperature (4 sec. at body temperature), and toxicity to human body seems low. For dental application, PTW was effective on infected models of human extracted tooth. Although PTW has many chemical components, respective chemical components in PTW were isolated by ion chromatography. In addition to peaks of H2O2, NO2- and NO3-, a specific peak was detected. and only this fraction had bactericidal activity. Purified active ingredient of PTW is the precursor of HOO, and further details will be discussed in the presentation. MEXT (15H03583, 23340176, 25108505). NCCE (23-A-15).

Phagocytosis, bacterial killing, and cytokine activation of circulating blood neutrophils in horses with severe equine asthma and control horses.

PubMed

Vanderstock, Johanne M; Lecours, Marie-Pier; Lavoie-Lamoureux, Annouck; Gottschalk, Marcelo; Segura, Mariela; Lavoie, Jean-Pierre; Jean, Daniel

2018-04-01

OBJECTIVE To evaluate in vitro phagocytosis and bactericidal activity of circulating blood neutrophils in horses with severe equine asthma and control horses and to determine whether circulating blood neutrophils in horses with severe equine asthma have an increase in expression of the proinflammatory cytokine tumor necrosis factor (TNF)-α and the chemokine interleukin (IL)-8 and a decrease in expression of the anti-inflammatory cytokine IL-10 in response to bacteria. ANIMALS 6 horses with severe equine asthma and 6 control horses. PROCEDURES Circulating blood neutrophils were isolated from horses with severe equine asthma and control horses. Phagocytosis was evaluated by use of flow cytometry. Bactericidal activity of circulating blood neutrophils was assessed by use of Streptococcus equi and Streptococcus zooepidemicus as targets, whereas the cytokine mRNA response was assessed by use of a quantitative PCR assay. RESULTS Circulating blood neutrophils from horses with severe equine asthma had significantly lower bactericidal activity toward S zooepidemicus but not toward S equi, compared with results for control horses. Phagocytosis and mRNA expression of TNF-α, IL-8, and IL-10 were not different between groups. CONCLUSIONS AND CLINCAL RELEVANCE Impairment of bactericidal activity of circulating blood neutrophils in horses with severe equine asthma could contribute to an increased susceptibility to infections.

Bactericidal activity of herbal volatile oil extracts against multidrug-resistant Acinetobacter baumannii.

PubMed

Intorasoot, Amornrat; Chornchoem, Piyaorn; Sookkhee, Siriwoot; Intorasoot, Sorasak

2017-01-01

The aim of the study is to investigate the antibacterial activity of 10 volatile oils extracted from medicinal plants, including galangal ( Alpinia galanga Linn.), ginger ( Zingiber officinale ), plai ( Zingiber cassumunar Roxb.), lime ( Citrus aurantifolia ), kaffir lime ( Citrus hystrix DC.), sweet basil ( Ocimum basilicum Linn.), tree basil ( Ocimum gratissimum ), lemongrass ( Cymbopogon citratus DC.), clove ( Syzygium aromaticum ), and cinnamon ( Cinnamomum verum ) against four standard strains of Staphylococcus aureus , Escherichia coli , Pseudomonas aeruginosa , Acinetobacter baumannii , and 30 clinical isolates of multidrug-resistant A. baumannii (MDR- A. baumannii ). Agar diffusion, minimum inhibitory concentration, and minimum bactericidal concentration (MBC) were employed for the determination of bactericidal activity of water distilled medicinal plants. Tea tree oil ( Melaleuca alternifolia ) was used as positive control in this study. The results indicated the volatile oil extracted from cinnamon exhibited potent antibacterial activity against the most common human pathogens, S. aureus , E. coli , P. aeruginosa , and A. baumannii . Most of volatile oil extracts were less effective against non-fermentative bacteria, P. aeruginosa . In addition, volatile oil extracted from cinnamon, clove, and tree basil possessed potent bactericidal activity against MDR- A. baumannii with MBC 90 of 0.5, 1, and 2 mg/mL, respectively. The volatile oil extracts would be useful as alternative natural product for the treatment of the most common human pathogens and MDR- A. baumannii infections.

An investigation of the bactericidal activity of chlorhexidine digluconateagainst multidrug-resistant hospital isolates.

PubMed

Ekizoğlu, Melike; Sağiroğlu, Meral; Kiliç, Ekrem; Hasçelik, Ayşe Gülşen

2016-04-19

Hospital infections are among the most prominent medical problems around the world. Using proper biocides in an appropriate way is critically important in overcoming this problem. Several reports have suggested that microorganisms may develop resistance or reduce their susceptibility to biocides, similar to the case with antibiotics. In this study we aimed to determine the antimicrobial activity of chlorhexidine digluconate against clinical isolates. The susceptibility of 120 hospital isolated strains of 7 bacterial genera against chlorhexidine digluconate was determined by agar dilution test, using minimum inhibitory concentration (MIC) values and the EN 1040 Basic Bactericidal Activity Test to determine the bactericidal activity. According to MIC values, Pseudomonas aeruginosa and Stenotrophomonas maltophilia were found to be less susceptible to chlorhexidine digluconate. Quantitative suspension test results showed that 4% chlorhexidine digluconate was effective against antibiotic resistant and susceptible bacteria after 5 min of contact time and can be safely used in our hospital. However, concentrations below 4% chlorhexidine digluconate caused a decrease in bactericidal activity, especially for Staphylococcus aureus and P. aeruginosa. It is crucial to use biocides at appropriate concentrations and to perform surveillance studies to trace resistance or low susceptibility patterns of S. aureus, P. aeruginosa, and other hospital isolates.

Bactericidal activity of electrolyzed acid water from solution containing sodium chloride at low concentration, in comparison with that at high concentration.

PubMed

Kiura, Hiromasa; Sano, Kouichi; Morimatsu, Shinichi; Nakano, Takashi; Morita, Chizuko; Yamaguchi, Masaki; Maeda, Toyoyuki; Katsuoka, Yoji

2002-05-01

Electrolyzed strong acid water (ESW) containing free chlorine at various concentrations is becoming to be available in clinical settings as a disinfectant. ESW is prepared by electrolysis of a NaCl solution, and has a corrosive activity against medical instruments. Although lower concentrations of NaCl and free chlorine are desired to eliminate corrosion, the germicidal effect of ESW with low NaCl and free-chlorine concentrations (ESW-L) has not been fully clarified. In this study, we demonstrated that ESW-L possesses bactericidal activity against Mycobacteria and spores of Bacillus subtilis. The effect was slightly weaker than that of ESW containing higher NaCl and free-chlorine concentrations (ESW-H), but acceptable as a disinfectant. To clarify the mechanism of the bactericidal activity, we investigated ESW-L-treated Pseudomonas aeruginosa by transmission electron microscopy, a bacterial enzyme assay and restriction fragment length polymorphism pattern (RFLP) assay. Since the bacterium, whose growth was completely inhibited by ESW-L, revealed the inactivation of cytoplasmic enzyme, blebs and breaks in its outer membrane and remained complete RFLP of DNA, damage of the outer membrane and inactivation of cytoplasmic enzyme are the important determinants of the bactericidal activity.

Acquisition of Complement Inhibitor Serine Protease Factor I and Its Cofactors C4b-Binding Protein and Factor H by Prevotella intermedia

PubMed Central

Malm, Sven; Jusko, Monika; Eick, Sigrun; Potempa, Jan; Riesbeck, Kristian; Blom, Anna M.

2012-01-01

Infection with the Gram-negative pathogen Prevotella intermedia gives rise to periodontitis and a growing number of studies implies an association of P. intermedia with rheumatoid arthritis. The serine protease Factor I (FI) is the central inhibitor of complement degrading complement components C3b and C4b in the presence of cofactors such as C4b-binding protein (C4BP) and Factor H (FH). Yet, the significance of complement inhibitor acquisition in P. intermedia infection and FI binding by Gram-negative pathogens has not been addressed. Here we show that P. intermedia isolates bound purified FI as well as FI directly from heat-inactivated human serum. FI bound to bacteria retained its serine protease activity as shown in degradation experiments with 125I-labeled C4b. Since FI requires cofactors for its activity we also investigated the binding of purified cofactors C4BP and FH and found acquisition of both proteins, which retained their activity in FI mediated degradation of C3b and C4b. We propose that FI binding by P. intermedia represents a new mechanism contributing to complement evasion by a Gram-negative bacterial pathogen associated with chronic diseases. PMID:22514678

Acquisition of complement inhibitor serine protease factor I and its cofactors C4b-binding protein and factor H by Prevotella intermedia.

PubMed

Malm, Sven; Jusko, Monika; Eick, Sigrun; Potempa, Jan; Riesbeck, Kristian; Blom, Anna M

2012-01-01

Infection with the Gram-negative pathogen Prevotella intermedia gives rise to periodontitis and a growing number of studies implies an association of P. intermedia with rheumatoid arthritis. The serine protease Factor I (FI) is the central inhibitor of complement degrading complement components C3b and C4b in the presence of cofactors such as C4b-binding protein (C4BP) and Factor H (FH). Yet, the significance of complement inhibitor acquisition in P. intermedia infection and FI binding by Gram-negative pathogens has not been addressed. Here we show that P. intermedia isolates bound purified FI as well as FI directly from heat-inactivated human serum. FI bound to bacteria retained its serine protease activity as shown in degradation experiments with (125)I-labeled C4b. Since FI requires cofactors for its activity we also investigated the binding of purified cofactors C4BP and FH and found acquisition of both proteins, which retained their activity in FI mediated degradation of C3b and C4b. We propose that FI binding by P. intermedia represents a new mechanism contributing to complement evasion by a Gram-negative bacterial pathogen associated with chronic diseases.

Subtle Variations in Surface Properties of Black Silicon Surfaces Influence the Degree of Bactericidal Efficiency

NASA Astrophysics Data System (ADS)

Bhadra, Chris M.; Werner, Marco; Baulin, Vladimir A.; Truong Khanh, Vi; Kobaisi, Mohammad Al; Nguyen, Song Ha; Balcytis, Armandas; Juodkazis, Saulius; Wang, James Y.; Mainwaring, David E.; Crawford, Russell J.; Ivanova, Elena P.

2018-06-01

One of the major challenges faced by the biomedical industry is the development of robust synthetic surfaces that can resist bacterial colonization. Much inspiration has been drawn recently from naturally occurring mechano-bactericidal surfaces such as the wings of cicada ( Psaltoda claripennis) and dragonfly ( Diplacodes bipunctata) species in fabricating their synthetic analogs. However, the bactericidal activity of nanostructured surfaces is observed in a particular range of parameters reflecting the geometry of nanostructures and surface wettability. Here, several of the nanometer-scale characteristics of black silicon (bSi) surfaces including the density and height of the nanopillars that have the potential to influence the bactericidal efficiency of these nanostructured surfaces have been investigated. The results provide important evidence that minor variations in the nanoarchitecture of substrata can substantially alter their performance as bactericidal surfaces.[Figure not available: see fulltext.

Binding of Soluble Yeast β-Glucan to Human Neutrophils and Monocytes is Complement-Dependent

PubMed Central

Bose, Nandita; Chan, Anissa S. H.; Guerrero, Faimola; Maristany, Carolyn M.; Qiu, Xiaohong; Walsh, Richard M.; Ertelt, Kathleen E.; Jonas, Adria Bykowski; Gorden, Keith B.; Dudney, Christine M.; Wurst, Lindsay R.; Danielson, Michael E.; Elmasry, Natalie; Magee, Andrew S.; Patchen, Myra L.; Vasilakos, John P.

2013-01-01

The immunomodulatory properties of yeast β-1,3/1,6 glucans are mediated through their ability to be recognized by human innate immune cells. While several studies have investigated binding of opsonized and unopsonized particulate β-glucans to human immune cells mainly via complement receptor 3 (CR3) or Dectin-1, few have focused on understanding the binding characteristics of soluble β-glucans. Using a well-characterized, pharmaceutical-grade, soluble yeast β-glucan, this study evaluated and characterized the binding of soluble β-glucan to human neutrophils and monocytes. The results demonstrated that soluble β-glucan bound to both human neutrophils and monocytes in a concentration-dependent and receptor-specific manner. Antibodies blocking the CD11b and CD18 chains of CR3 significantly inhibited binding to both cell types, establishing CR3 as the key receptor recognizing the soluble β-glucan in these cells. Binding of soluble β-glucan to human neutrophils and monocytes required serum and was also dependent on incubation time and temperature, strongly suggesting that binding was complement-mediated. Indeed, binding was reduced in heat-inactivated serum, or in serum treated with methylamine or in serum reacted with the C3-specific inhibitor compstatin. Opsonization of soluble β-glucan was demonstrated by detection of iC3b, the complement opsonin on β-glucan-bound cells, as well as by the direct binding of iC3b to β-glucan in the absence of cells. Binding of β-glucan to cells was partially inhibited by blockade of the alternative pathway of complement, suggesting that the C3 activation amplification step mediated by this pathway also contributed to binding. PMID:23964276

Exploiting a novel conformational switch to control innate immunity mediated by complement protein C3a.

PubMed

Lohman, Rink-Jan; Hamidon, Johan K; Reid, Robert C; Rowley, Jessica A; Yau, Mei-Kwan; Halili, Maria A; Nielsen, Daniel S; Lim, Junxian; Wu, Kai-Chen; Loh, Zhixuan; Do, Anh; Suen, Jacky Y; Iyer, Abishek; Fairlie, David P

2017-08-24

Complement C3a is an important protein in innate and adaptive immunity, but its specific roles in vivo remain uncertain because C3a degrades rapidly to form the C3a-desArg protein, which does not bind to the C3a receptor and is indistinguishable from C3a using antibodies. Here we develop the most potent, stable and highly selective small molecule modulators of C3a receptor, using a heterocyclic hinge to switch between agonist and antagonist ligand conformations. This enables characterization of C3 areceptor-selective pro- vs. anti-inflammatory actions in human mast cells and macrophages, and in rats. A C3a receptor-selective agonist induces acute rat paw inflammation by first degranulating mast cells before activating macrophages and neutrophils. An orally administered C3a receptor-selective antagonist inhibits mast cell degranulation, thereby blocking recruitment and activation of macrophages and neutrophils, expression of inflammatory mediators and inflammation in a rat paw edema model. These novel tools reveal the mechanism of C3a-induced inflammation and provide new insights to complement-based medicines.Complement C3a is an important protein in innate and adaptive immunity, but its roles in vivo are unclear. Here the authors develop novel chemical agonists and antagonists for the C3a receptor, and show that they modulate mast cell degranulation and inflammation in a rat paw edema model.

Immunization with recombinant truncated Neisseria meningitidis-Macrophage Infectivity Potentiator (rT-Nm-MIP) protein induces murine antibodies that are cross-reactive and bactericidal for Neisseria gonorrhoeae.

PubMed

Humbert, María Victoria; Christodoulides, Myron

2018-05-23

Neisseria meningitidis (Nm) and N. gonorrhoeae (Ng) express a Macrophage Infectivity Potentiator (MIP, NMB1567/NEIS1487) protein in their outer membrane (OM). In this study, we prepared independent batches of liposomes (n = 3) and liposomes + MonoPhosphoryl Lipid A (MPLA) (n = 3) containing recombinant truncated Nm-MIP protein encoded by Allele 2 (rT-Nm-MIP, amino acids 22-142), and used these to immunize mice. We tested the hypothesis that independent vaccine batches showed similar antigenicity, and that antisera could recognise both meningococcal and gonococcal MIP and induce cross-species bactericidal activity. The different batches of M2 rT-Nm-MIP-liposomes ± MPLA showed no significant (P > 0.05) batch-to-batch variation in antigenicity. Anti-rT-Nm-MIP sera reacted equally and specifically with Nm-MIP and Ng-MIP in OM and on live bacterial cell surfaces. Specificity was shown by no antiserum reactivity with Δmip bacteria. Using human complement/serum bactericidal assays, anti-M2 rT-Nm-MIP sera killed homologous meningococcal serogroup B (MenB) strains (median titres of 32-64 for anti-rT-Nm-MIP-liposome sera; 128-256 for anti-rT-Nm-MIP-liposome + MPLA sera) and heterologous M1 protein-expressing MenB strains (titres of 64 for anti rT-Nm-MIP-liposome sera; 128-256 for anti-rT-Nm-MIP-liposome + MPLA sera). Low-level killing (P < 0.05) was observed for a MenB isolate expressing M7 protein (titres 4-8), but MenB strains expressing M6 protein were not killed (titre < 4-8). Killing (P < 0.05) was observed against MenC and MenW bacteria expressing homologous M2 protein (titres of 8-16) but not against MenA or MenY bacteria (titres < 4-8). Antisera to M2 rT-Nm-MIP showed significant (P < 0.05) cross-bactericidal activity against gonococcal strain P9-17 (expressing M35 Ng-MIP, titres of 64-512) and strain 12CFX_T_003 (expressing M10 Ng-MIP, titres 8-16) but not against FA1090 (expressing M8 Ng-MIP). As an alternative to producing recombinant protein, we engineered successfully the Nm-OM to express M2 Truncated-Nm-MIP, but lipooligosaccharide-extraction with Na-DOC was contra-indicated. Our data suggest that a multi-component vaccine containing a select number of Nm- and Ng-MIP type proteins would be required to provide broad coverage of both pathogens. Copyright © 2018. Published by Elsevier Ltd.

Antibody mediated rejection associated with complement factor h-related protein 3/1 deficiency successfully treated with eculizumab.

PubMed

Noone, D; Al-Matrafi, J; Tinckam, K; Zipfel, P F; Herzenberg, A M; Thorner, P S; Pluthero, F G; Kahr, W H A; Filler, G; Hebert, D; Harvey, E; Licht, C

2012-09-01

Antibody mediated rejection (AMR) activates the classical complement pathway and can be detrimental to graft survival. AMR can be accompanied by thrombotic microangiopathy (TMA). Eculizumab, a monoclonal C5 antibody prevents induction of the terminal complement cascade (TCC) and has recently emerged as a therapeutic option for AMR. We present a highly sensitized 13-year-old female with end-stage kidney disease secondary to spina bifida-associated reflux nephropathy, who developed severe steroid-, ATG- and plasmapheresis-resistant AMR with TMA 1 week post second kidney transplant despite previous desensitization therapy with immunoglobulin infusions. Eculizumab rescue therapy resulted in a dramatic improvement in biochemical (C3; creatinine) and hematological (platelets) parameters within 6 days. The patient was proven to be deficient in complement Factor H-related protein 3/1 (CFHR3/1), a plasma protein that regulates the complement cascade at the level of C5 conversion and has been involved in the pathogenesis of atypical hemolytic uremic syndrome caused by CFH autoantibodies (DEAP-HUS). CFHR1 deficiency may have worsened the severe clinical progression of AMR and possibly contributed to the development of donor-specific antibodies. Thus, screening for CFHR3/1 deficiency should be considered in patients with severe AMR associated with TMA. © Copyright 2012 The American Society of Transplantation and the American Society of Transplant Surgeons.

Bactericidal Effect of Photolysis of H2O2 in Combination with Sonolysis of Water via Hydroxyl Radical Generation.

PubMed

Sheng, Hong; Nakamura, Keisuke; Kanno, Taro; Sasaki, Keiichi; Niwano, Yoshimi

2015-01-01

The bactericidal effect of hydroxyl radical (·OH) generated by combination of photolysis of hydrogen peroxide (H2O2) and sonolysis of water was examined under the condition in which the yield of ·OH increased additively when H2O2 aqueous solution was concomitantly irradiated with laser and ultrasound. The suspension of Staphylococcus aureus mixed with the different concentrations of H2O2 was irradiated simultaneously with a laser light (wavelength: 405 nm, irradiance: 46 and 91 mW/cm2) and ultrasound (power: 30 w, frequency: 1.65 MHz) at 20 ± 1°C of the water bulk temperature for 2 min. The combination of laser and ultrasound irradiation significantly reduced the viable bacterial count in comparison with the laser irradiation of H2O2 alone. By contrast, the ultrasound irradiation alone exerted almost no bactericidal effect. These results suggested that the combination effect of photolysis of H2O2 and sonolysis of water on bactericidal activity was synergistic. A multi-way analysis of variance also revealed that the interaction of H2O2 concentration, laser power and ultrasound irradiation significantly affected the bactericidal activity. Since the result of oxidative DNA damage evaluation demonstrated that the combination of laser and ultrasound irradiation significantly induced oxidative damage of bacterial DNA in comparison with the laser irradiation of H2O2 alone, it was suggested that the combination effect of photolysis of H2O2 and sonolysis of water on bactericidal activity would be exerted via oxidative damage of cellular components such as DNA.

Dengue virus induces increased activity of the complement alternative pathway in infected cells.

PubMed

Cabezas, Sheila; Bracho, Gustavo; L Aloia, Amanda; Adamson, Penelope J; Bonder, Claudine S; Smith, Justine R; Gordon, David L; Carr, Jillian M

2018-05-09

Severe dengue virus (DENV) infection is associated with overactivity of the complement alternative pathway (AP) in patient studies. Here, the molecular changes in components of the AP during DENV infection in vitro are investigated. mRNA for factor H (FH) a major negative regulator of the AP, is significantly increased in DENV-infected endothelial cells (EC) and macrophages but in contrast production of extracellular FH protein is not. This discord is not seen for the AP activator, factor B (FB), with DENV induction of both FB mRNA and protein, nor with Toll-like receptor 3 or 4 stimulation of EC and macrophages, which induces both FH and FB mRNA and protein. Surface bound and intracellular FH protein is however induced by DENV, but only in DENV antigen-positive cells, while in two other DENV-susceptible immortalised cell lines (ARPE-19 and HREC) FH protein is induced both intracellularly and extracellularly by DENV infection. Regardless of the cell type, there is an imbalance in AP components and an increase in markers of complement AP activity associated with DENV-infected cells - with lower FH relative to FB protein, increased ability to promote AP-mediated lytic activity and increased deposition of complement component C3b on the surface of DENV-infected cells. For EC in particular, these changes are predicted to result in higher complement activity in the local cellular microenvironment, with the potential to induce functional changes that may result in increased vascular permeability, a hallmark of dengue disease. IMPORTANCE Dengue virus (DENV) is a significant human viral pathogen with global medical and economic impact. DENV may cause serious and life-threatening disease with increased vascular permeability and plasma leakage. The pathogenic mechanisms underlying these features remain unclear; however overactivity of the complement alternative pathway has been suggested to play a role. In this study we investigate the molecular events that may be responsible for this observed alternative pathway overactivity and provide novel findings of changes in the complement system in response to DENV infection in primary cell types that are a major target for DENV infection (macrophages) and pathogenesis (endothelial cells) in vivo Our results suggest a new dimension of cellular events that may influence endothelial cell barrier function during DENV infection that could expand strategies for developing therapeutics to prevent or control DENV-mediated vascular disease. Copyright © 2018 American Society for Microbiology.

Synthesis of Silver nanoparticles (AgNPs) with Antibacterial Activity

NASA Astrophysics Data System (ADS)

Campillo Gloria, E.; Ederley, Vélez; Gladis, Morales; César, Hincapié; Jaime, Osorio; Oscar, Arnache; Uribe José, Ignacio; Franklin, Jaramillo

2017-06-01

The synthesis of nanomaterials is currently one of the most active in nanoscience branches; especially those help improve the human quality life. Silver nanoparticles (AgNPs) are an example of this as it is known to have inhibitory and bactericidal effects. In this work, we report the synthesis of silver nanoparticles by chemical reduction method of silver nitrate (AgNO3) from aqueous solution, using a mix of polivinyl pyrrolidone (PVP) - Aloe Vera as reducing agent and for stabilization and control of particle size. Silver nanoparticles obtained were characterized by Scanning Electron Microscopy (SEM), UV-visible spectroscopy and measurements using Zetasizer Nano ZS were applied to size estimation. The existence of surface plasmon resonance peak at λmax ~ 420 nm is evidence of silver nanoparticles formation. It was possible to standardize an appropriate protocol for the evaluation of bactericidal activity of the nanoparticles, for mesophilic microorganisms. Bactericidal activity above 90% against these kinds of bacteria was demonstrated.

Antibacterial Activity of Cinoxacin In Vitro

PubMed Central

Giamarellou, Helen; Jackson, George G.

1975-01-01

Cinoxacin is a new synthetic compound similar chemically and in antimicrobial activity to oxolonic acid and nalidixic acid. It is most effective against Escherichia coli and Proteus mirabilis, but at concentrations expected in the urine it is inhibitory for all species of Enterobacteriaceae. Relative to nalidixic acid, cinoxacin has slightly greater inhibitory and bactericidal activity, less inoculum effect probably due to less heterogeneity in the susceptibility of bacterial cells, and less inhibition by high concentrations of serum protein. Both drugs are more active in an acid than an alkaline medium. Glucose can specifically antagonize the inhibitory effect against P. mirabilis. In urine the bactericidal rate and effect are decreased. Resistance to cinoxacin can be developed quickly by serial transfers in vitro. Some nonresistant organisms remained viable in bactericidal drug concentrations. The in vivo importance of the favorable features of cinoxacin must be determined by clinical trials. PMID:1096811

Evaluation of bactericidal effects of low-temperature nitrogen gas plasma towards application to short-time sterilization.

PubMed

Kawamura, Kumiko; Sakuma, Ayaka; Nakamura, Yuka; Oguri, Tomoko; Sato, Natsumi; Kido, Nobuo

2012-07-01

To develop a novel low-temperature plasma sterilizer using pure N(2) gas as a plasma source, we evaluated bactericidal ability of a prototype apparatus provided by NGK Insulators. After determination of the sterilizing conditions without the cold spots, the D value of the BI of Geobacillus stearothermophilus endospores on the filter paper was determined as 1.9 min. However, the inactivation efficiency of BI carrying the same endospores on SUS varied to some extent, suggesting that the bactericidal effect might vary by materials of sterilized instruments. Staphylococcus aureus and Escherichia coli were also exposed to the N(2) gas plasma and confirmed to be inactivated within 30 min. Through the evaluation of bactericidal efficiency in a sterilization bag, we concluded that the UV photons in the plasma and the high-voltage pulse to generate the gas plasma were not concerned with the bactericidal effect of the N(2) gas plasma. Bactericidal effect might be exhibited by activated nitrogen atoms or molecular radicals. © 2012 The Societies and Blackwell Publishing Asia Pty Ltd.

Persistence of the immune response after MenACWY-CRM vaccination and response to a booster dose, in adolescents, children and infants

PubMed Central

Baxter, Roger; Keshavan, Pavitra; Welsch, Jo Anne; Han, Linda; Smolenov, Igor

2016-01-01

ABSTRACT Persistence of bactericidal antibodies following vaccination is extremely important for protection against invasive meningococcal disease, given the epidemiology and rapid progression of meningococcal infection. We present an analysis of antibody persistence and booster response to MenACWY-CRM, in adolescents, children and infants, from 7 clinical studies. Immunogenicity was assessed using the serum bactericidal assay with both human and rabbit complement. Post-vaccination hSBA titers were high, with an age- and serogroup-specific decline in titers up to 1 y and stable levels up to 5 y The waning of hSBA titers over time was more pronounced among infants and toddlers and the greatest for serogroup A. However, rSBA titers against serogroup A were consistently higher and showed little decline over time, suggesting that protection against this serogroup may be sustained. A single booster dose of MenACWY-CRM administered at 3 to 5 y induced a robust immune response in all age groups. PMID:26829877

Conjugation Approach To Produce a Staphylococcus aureus Synbody with Activity in Serum.

PubMed

Lainson, John C; Fuenmayor, Mariana Ferrer; Johnston, Stephen Albert; Diehnelt, Chris W

2015-10-21

Synbodies show promise as a new class of synthetic antibiotics. Here, we explore improvements in their activity and production through conjugation chemistry. Maleimide conjugation is a widely used conjugation strategy due to its high yield, selectivity, and low cost. We used this strategy to conjugate two antibacterial peptides to produce a bivalent antibacterial peptide, called a synbody that has bactericidal activity against methicillin resistant Staphylococcus aureus (MRSA). The synbody was prepared by conjugation of a partially d-amino acid substituted synthetic antibacterial peptide to a bis-maleimide scaffold. The synbody slowly degrades in serum, but also undergoes exchange reactions with other serum proteins, such as albumin. Therefore, we hydrolyzed the thiosuccinimide ring using a mild hydrolysis protocol to produce a new synbody with similar bactericidal activity. The synbody was now resistant to exchange reactions and maintained bactericidal activity in serum for 2 h. This work demonstrates that low-cost maleimide coupling can be used to produce antibacterial peptide conjugates with activity in serum.

Adding Selectivity to Antimicrobial Peptides: Rational Design of a Multidomain Peptide against Pseudomonas spp.

PubMed Central

Eckert, Randal; Qi, Fengxia; Yarbrough, Daniel K.; He, Jian; Anderson, Maxwell H.; Shi, Wenyuan

2006-01-01

Currently available antimicrobials exhibit broad killing with regard to bacterial genera and species. Indiscriminate killing of microbes by these conventional antibiotics can disrupt the ecological balance of the indigenous microbial flora, often resulting in negative clinical consequences. Species-specific antimicrobials capable of precisely targeting pathogenic bacteria without damaging benign microorganisms provide a means of avoiding this problem. In this communication, we report the successful creation of the first synthetic, target-specific antimicrobial peptide, G10KHc, via addition of a rationally designed Pseudomonas-specific targeting moiety (KH) to a generally killing peptide (novispirin G10). The resulting chimeric peptide showed enhanced bactericidal activity and faster killing kinetics against Pseudomonas spp. than G10 alone. The enhanced killing activities are due to increased binding and penetration of the outer membrane of Pseudomonas sp. cells. These properties were not observed in tests of untargeted bacterial species, and this specificity allowed G10KHc to selectively eliminate Pseudomonas spp. from mixed cultures. This work lays a foundation for generating target-specific “smart” antimicrobials to complement currently available conventional antibiotics. PMID:16569868

Characterization of the extracellular bactericidal factors of rat alveolar lining material.

PubMed Central

Coonrod, J D; Lester, R L; Hsu, L C

1984-01-01

The surfactant fraction (55,000-g pellet) of leukocyte-free rat bronchoalveolar lavage fluid contains factors that rapidly kill and lyse pneumococci. These factors were purified and identified biochemically by using a quantitative bactericidal test to monitor fractionation procedures. 91% of the antipneumococcal activity of rat surfactant was recovered in chloroform after extraction of rat surfactant with chloroform-methanol (Bligh-Dyer procedure). After chromatography on silicic acid with chloroform, acetone, and methanol, all detectable antibacterial activity (approximately 80% of the initial activity) eluted with the neutral lipids in chloroform. When rechromatographed on silicic acid with hexane, hexane-chloroform, and chloroform, the antibacterial activity eluted with FFA. Thin-layer chromatography (TLC) established that the antibacterial activity was confined to the FFA fraction. Gas-liquid chromatography showed that the fatty acid fraction contained a mixture of long-chain FFA (C12 to C22) of which 66.7% were saturated and 32.4% were unsaturated. The quantity of TLC-purified FFA needed to kill 50% of 10(8) pneumococci under standardized conditions (one bactericidal unit) was 10.6 +/- 0.5 micrograms. Purified FFA acted as detergents, causing release of [3H]choline from pneumococcal cell walls and increased bacterial cell membrane permeability, evidenced by rapid unloading of 3-O-[3H]methyl-D-glucose. FFA acting as detergents appear to account for the bactericidal and bacteriolytic activity of rat pulmonary surfactant for pneumococci. PMID:6548228

Proinflammatory Activity of a Cecropin-Like Antibacterial Peptide from Helicobacter pylori

PubMed Central

Bylund, Johan; Christophe, Thierry; Boulay, Francois; Nyström, Thomas; Karlsson, Anna; Dahlgren, Claes

2001-01-01

Helicobacter pylori, the bacterial pathogen associated with gastritis and peptic ulcers, is highly successful in establishing infection in the human gastric mucosa, a process typically associated with massive infiltration of inflammatory cells. Colonization of the mucosa is suggested to be facilitated by H. pylori-produced cecropin-like peptides with antibacterial properties, giving the microbe a competitive advantage over other bacteria. We show that a cecropin-like antibacterial peptide from H. pylori, Hp(2-20), not only has a potent bactericidal effect but also induces proinflammatory activities in human neutrophils, e.g., upregulation of integrins (Mac-1), induction of chemotaxis, and activation of the oxygen radical producing NADPH-oxidase. Furthermore, we show that these effects are mediated through binding of Hp(2-20) to the promiscuous, G-protein-linked lipoxin A4 receptor–formyl peptide-like receptor 1. PMID:11353614

Preliminary flight prototype potable water bactericide system

NASA Technical Reports Server (NTRS)

Jasionowski, W. J.; Allen, E. T.

1973-01-01

The development, design, and testing of a preliminary flight prototype potable water bactericide system are described. The system is an assembly of upgraded canisters composed of: (1) A biological filter; (2) an activated charcoal and ion exchange resin canister; (3) a silver chloride canister, (4) a deionizer, (5) a silver bromide canister with a partial bypass, and (6) mock-up instrumentation and circuitry. The system exhibited bactericidal activity against 10 to the 9th power Pseudomonas aeruginosa and/or Type IIIa, and reduced Bacillus subtilis by up to 5 orders of magnitude in 24 hours at ambient temperatures with a 1 ppm silver ion dose. Four efficacy tests were performed with a AgBr canister dosing anticipated fuel cell water. Tests show that a 0.05 ppm silver ion dose was bactericidal against 3 plus or minus 1 x 10 to the 9th power (5 plus or minus 1 x 10,000/ml Pseudomonas aeruginosa and/or Type IIIa in 15 minutes or less.

Susceptibility of Escherichia coli to Bactericidal Action of Lactoperoxidase, Peroxide, and Iodide or Thiocyanate

PubMed Central

Thomas, Edwin L.; Aune, Thomas M.

1978-01-01

The bactericidal action that results from lactoperoxidase-catalyzed oxidation of iodide or thiocyanate was studied, using Escherichia coli as the test organism. The susceptibility of intact cells to bactericidal action was compared with that of cells with altered cell envelopes. Exposure to ethylenediaminetetraacetic acid, to lysozyme and ethylenediaminetetraacetic acid, or to osmotic shock were used to alter the cell envelope. Bactericidal action was greatly increased when the cells were exposed to the lactoperoxidase-peroxide-iodide system at low temperatures, low cell density, or after alteration of the cell envelope. When thiocyanate was substituted for iodide, bactericidal activity was observed only at low cell density or after osmotic shock. Low temperature and low cell density lowered the rate of destruction of peroxide by the bacteria. Therefore, competition for peroxide between the bacteria and lactoperoxidase may influence the extent of bactericidal action. Alteration of the cell envelope had only a small effect on the rate of destruction of peroxide. Instead, the increased susceptibility of these altered cells suggested that bactericidal action required permeation of a reagent through the cell envelope. In addition to altering the cell envelope, these procedures partly depleted cells of oxidizable substrates and sulfhydryl components. Adding an oxidizable substrate did not decrease the susceptibility of the altered cells. On the other hand, mild reducing agents such as sulfhydryl compounds did partly reverse bactericidal action when added after exposure of cells to the peroxidase systems. These studies indicate that alteration of the metabolism, structure, or composition of bacterial cells can greatly increase their susceptibility to peroxidase bactericidal action. PMID:348097

Influence of IgG Subclass on Human Antimannan Antibody-Mediated Resistance to Hematogenously Disseminated Candidiasis in Mice

PubMed Central

Nishiya, Casey T.; Boxx, Gayle M.; Robison, Kerry; Itatani, Carol; Kozel, Thomas R.

2015-01-01

Candida albicans is a yeast-like pathogen and can cause life-threatening systemic candidiasis. Its cell surface is enriched with mannan that is resistant to complement activation. Previously, we developed the recombinant human IgG1 antimannan antibody M1g1. M1g1 was found to promote complement activation and phagocytosis and protect mice from systemic candidiasis. Here, we evaluate the influence of IgG subclass on antimannan antibody-mediated protection. Three IgG subclass variants of M1g1 were constructed: M1g2, M1g3, and M1g4. The IgG subclass identity for each variant was confirmed with DNA sequence and subclass-specific antibodies. These variants contain identical M1 Fabs and exhibited similar binding affinities for C. albicans yeast and purified mannan. Yeast cells and hyphae recovered from the kidney of antibody-treated mice with systemic candidiasis showed uniform binding of each variant, indicating constitutive expression of the M1 epitope and antibody opsonization in the kidney. All variants promoted deposition of both murine and human C3 onto the yeast cell surface, with M1g4 showing delayed activation, as determined by flow cytometry and immunofluorescence microscopy. M1g4-mediated complement activation was found to be associated with its M1 Fab that activates the alternative pathway in an Fc-independent manner. Treatment with each subclass variant extended the survival of mice with systemic candidiasis (P < 0.001). However, treatment with M1g1, M1g3, or M1g4, but not with M1g2, also reduced the kidney fungal burden (P < 0.001). Thus, the role of human antimannan antibody in host resistance to systemic candidiasis is influenced by its IgG subclass. PMID:26573736

Effects of rearing temperature on immune functions in sockeye salmon (Oncorhynchus nerka)

USGS Publications Warehouse

Alcorn, S.W.; Murray, A.L.; Pascho, R.J.

2002-01-01

To determine if the defences of sockeye salmon (Oncorhynchus nerka) raised in captivity are affected by the rearing temperature or their life-cycle stage, various indices of the humoral and cellular immune functions were measured in fish reared at either 8 or 12??C for their entire life-cycle. Measures of humoral immunity included the commonly used haematological parameters, as well as measurements of complement, and lysozyme activity. Cellular assays quantified the ability of macrophages from the anterior kidney to phagocytise Staphylococcus aureus cells, or the activities of certain bactericidal systems of those cells. The T-dependent antibody response to a recombinant 57 kDa protein of Renibacterium salmoninarum was used to quantify the specific immune response. Fish were sampled during the spring and fall of their second, third and fourth years, corresponding to a period that began just before smolting and ended at sexual maturation. Fish reared at 8??C tended to have a greater percentage of phagocytic kidney macrophages during the first 2 years of sampling than the fish reared at 12??C. During the last half of the study the complement activity of the fish reared at 8??C was greater than that of the 12??C fish. Conversely, a greater proportion of the blood leucocytes were lymphocytes in fish reared at 12??C compared to the fish reared at 8??C. Fish reared at 12??C also produced a greater antibody response than those reared at 8??C. Results suggested that the immune apparatus of sockeye salmon reared at 8??C relied more heavily on the non-specific immune response, while the specific immune response was used to a greater extent when the fish were reared at 12??C. Although a seasonal effect was not detected in any of the indices measured, varying effects were observed in some measurements during sexual maturation of fish in both temperature groups. At that time there were dramatic decreases in complement activity and lymphocyte numbers. This study was unique in its scope because it was the first quantitative assessment of salmon immune functions for an entire life-cycle. ?? 2002 Elsevier Science Ltd.

Antibacterial activity of DLC films containing TiO2 nanoparticles.

PubMed

Marciano, F R; Lima-Oliveira, D A; Da-Silva, N S; Diniz, A V; Corat, E J; Trava-Airoldi, V J

2009-12-01

Diamond-like carbon (DLC) films have been the focus of extensive research in recent years due to their potential applications as surface coatings on biomedical devices. Titanium dioxide (TiO2) in the anatase crystalline form is a strong bactericidal agent when exposed to near-UV light. In this work we investigate the bactericidal activity of DLC films containing TiO2 nanoparticles. The films were grown on 316L stainless-steel substrates from a dispersion of TiO2 in hexane using plasma-enhanced chemical vapor deposition. The composition, bonding structure, surface energy, stress, and surface roughness of these films were also evaluated. The antibacterial tests were performed against Escherichia coli (E. coli) and the results were compared to the bacterial adhesion force to the studied surfaces. The presence of TiO2 in DLC bulk was confirmed by Raman spectroscopy. As TiO2 content increased, I(D)/I(G) ratio, hydrogen content, and roughness also increased; the films became more hydrophilic, with higher surface free energy and the interfacial energy of bacteria adhesion decreased. Experimental results show that TiO2 increased DLC bactericidal activity. Pure DLC films were thermodynamically unfavorable to bacterial adhesion. However, the chemical interaction between the E. coli and the studied films increased for the films with higher TiO2 concentration. As TiO2 bactericidal activity starts its action by oxidative damage to the bacteria wall, a decrease in the interfacial energy of bacteria adhesion causes an increase in the chemical interaction between E. coli and the films, which is an additional factor for the increasing bactericidal activity. From these results, DLC with TiO2 nanoparticles can be useful for producing coatings with antibacterial properties.

Peptide fragments of a beta-defensin derivative with potent bactericidal activity.

PubMed

Reynolds, Natalie L; De Cecco, Martin; Taylor, Karen; Stanton, Chloe; Kilanowski, Fiona; Kalapothakis, Jason; Seo, Emily; Uhrin, Dusan; Campopiano, Dominic; Govan, John; Macmillan, Derek; Barran, Perdita; Dorin, Julia R

2010-05-01

Beta-defensins are known to be both antimicrobial and able to chemoattract various immune cells. Although the sequences of paralogous genes are not highly conserved, the core defensin structure is retained. Defb14-1C(V) has bactericidal activity similar to that of its parent peptide (murine beta-defensin Defb14) despite all but one of the canonical six cysteines being replaced with alanines. The 23-amino-acid N-terminal half of Defb14-1C(V) is a potent antimicrobial while the C-terminal half is not. Here, we use a library of peptide derivatives to demonstrate that the antimicrobial activity can be localized to a particular region. Overlapping fragments of the N-terminal region were tested for their ability to kill Gram-positive and Gram-negative bacteria. We demonstrate that the most N-terminal fragments (amino acids 1 to 10 and 6 to 17) are potent antimicrobials against Gram-negative bacteria whereas fragments based on sequence more C terminal than amino acid 13 have very poor activity against both Gram-positive and -negative types. We further test a series of N-terminal deletion peptides in both their monomeric and dimeric forms. We find that bactericidal activity is lost against both Gram types as the deletion region increases, with the point at which this occurs varying between bacterial strains. The dimeric form of the peptides is more resistant to the peptide deletions, but this is not due just to increased charge. Our results indicate that the primary sequence, together with structure, is essential in the bactericidal action of this beta-defensin derivative peptide and importantly identifies a short fragment from the peptide that is a potent bactericide.

Peptide Fragments of a β-Defensin Derivative with Potent Bactericidal Activity ▿

PubMed Central

Reynolds, Natalie L.; De Cecco, Martin; Taylor, Karen; Stanton, Chloe; Kilanowski, Fiona; Kalapothakis, Jason; Seo, Emily; Uhrin, Dusan; Campopiano, Dominic; Govan, John; Macmillan, Derek; Barran, Perdita; Dorin, Julia R.

2010-01-01

β-Defensins are known to be both antimicrobial and able to chemoattract various immune cells. Although the sequences of paralogous genes are not highly conserved, the core defensin structure is retained. Defb14-1CV has bactericidal activity similar to that of its parent peptide (murine β-defensin Defb14) despite all but one of the canonical six cysteines being replaced with alanines. The 23-amino-acid N-terminal half of Defb14-1CV is a potent antimicrobial while the C-terminal half is not. Here, we use a library of peptide derivatives to demonstrate that the antimicrobial activity can be localized to a particular region. Overlapping fragments of the N-terminal region were tested for their ability to kill Gram-positive and Gram-negative bacteria. We demonstrate that the most N-terminal fragments (amino acids 1 to 10 and 6 to 17) are potent antimicrobials against Gram-negative bacteria whereas fragments based on sequence more C terminal than amino acid 13 have very poor activity against both Gram-positive and -negative types. We further test a series of N-terminal deletion peptides in both their monomeric and dimeric forms. We find that bactericidal activity is lost against both Gram types as the deletion region increases, with the point at which this occurs varying between bacterial strains. The dimeric form of the peptides is more resistant to the peptide deletions, but this is not due just to increased charge. Our results indicate that the primary sequence, together with structure, is essential in the bactericidal action of this β-defensin derivative peptide and importantly identifies a short fragment from the peptide that is a potent bactericide. PMID:20176896

Polyethylenimine mediated silver nanoparticle-decorated magnetic graphene as a promising photothermal antibacterial agent

NASA Astrophysics Data System (ADS)

Wang, Ning; Hu, Bo; Chen, Ming-Li; Wang, Jian-Hua

2015-05-01

A novel bactericidal material, Ag@rGO-Fe3O4-PEI composite is prepared by in situ growth of silver nanoparticles onto the polyethylenimine (PEI)-mediated magnetic reduced graphene oxide (GO). The antibacterial performances of the composite are investigated by using the gram-negative bacteria Escherichia coli O157:H7 (E. coli O157:H7) as a model. The results indicate that the Ag@rGO-Fe3O4-PEI composite exhibits excellent antibacterial performance against E. coli O157:H7, with an antibacterial performance superior to those for the ever-reported photothermal materials. The bactericidal capability or the inhibition capability for bacteria growth is found to depend on the dosage of the Ag@rGO-Fe3O4-PEI and Ag/rGO-Fe3O4-PEI mass ratio within a certain range. By using a dosage of 0.1 μg mL-1, a killing rate of 99.9% is achieved for the E. coli O157:H7 (1 × 107 cfu mL-1) under a 0.5 min NIR laser irradiation (785 nm/50 mW cm-2). In addition, a minimum bactericidal concentration (MBC) of 0.100 μg mL-1 is achieved under near infrared (NIR) laser irradiation for 10 min, for which case there is absolutely no colony of E. coli O157:H7 found in the broth agar plate.

Virulence Associated Gene 8 of Bordetella pertussis Enhances Contact System Activity by Inhibiting the Regulatory Function of Complement Regulator C1 Inhibitor

PubMed Central

Hovingh, Elise S.; de Maat, Steven; Cloherty, Alexandra P. M.; Johnson, Steven; Pinelli, Elena; Maas, Coen; Jongerius, Ilse

2018-01-01

Bordetella pertussis is a Gram-negative bacterium and the causative agent of whooping cough. Whooping cough is currently re-emerging worldwide and, therefore, still poses a continuous global health threat. B. pertussis expresses several virulence factors that play a role in evading the human immune response. One of these virulence factors is virulence associated gene 8 (Vag8). Vag8 is a complement evasion molecule that mediates its effects by binding to the complement regulator C1 inhibitor (C1-INH). This regulatory protein is a fluid phase serine protease that controls proenzyme activation and enzyme activity of not only the complement system but also the contact system. Activation of the contact system results in the generation of bradykinin, a pro-inflammatory peptide. Here, the activation of the contact system by B. pertussis was explored. We demonstrate that recombinant as well as endogenous Vag8 enhanced contact system activity by binding C1-INH and attenuating its inhibitory function. Moreover, we show that B. pertussis itself is able to activate the contact system. This activation was dependent on Vag8 production as a Vag8 knockout B. pertussis strain was unable to activate the contact system. These findings show a previously overlooked interaction between the contact system and the respiratory pathogen B. pertussis. Activation of the contact system by B. pertussis may contribute to its pathogenicity and virulence. PMID:29915576

Virulence Associated Gene 8 of Bordetella pertussis Enhances Contact System Activity by Inhibiting the Regulatory Function of Complement Regulator C1 Inhibitor.

PubMed

Hovingh, Elise S; de Maat, Steven; Cloherty, Alexandra P M; Johnson, Steven; Pinelli, Elena; Maas, Coen; Jongerius, Ilse

2018-01-01

Bordetella pertussis is a Gram-negative bacterium and the causative agent of whooping cough. Whooping cough is currently re-emerging worldwide and, therefore, still poses a continuous global health threat. B. pertussis expresses several virulence factors that play a role in evading the human immune response. One of these virulence factors is virulence associated gene 8 (Vag8). Vag8 is a complement evasion molecule that mediates its effects by binding to the complement regulator C1 inhibitor (C1-INH). This regulatory protein is a fluid phase serine protease that controls proenzyme activation and enzyme activity of not only the complement system but also the contact system. Activation of the contact system results in the generation of bradykinin, a pro-inflammatory peptide. Here, the activation of the contact system by B. pertussis was explored. We demonstrate that recombinant as well as endogenous Vag8 enhanced contact system activity by binding C1-INH and attenuating its inhibitory function. Moreover, we show that B. pertussis itself is able to activate the contact system. This activation was dependent on Vag8 production as a Vag8 knockout B. pertussis strain was unable to activate the contact system. These findings show a previously overlooked interaction between the contact system and the respiratory pathogen B. pertussis . Activation of the contact system by B. pertussis may contribute to its pathogenicity and virulence.

Formation, clearance, deposition, pathogenicity, and identification of biopharmaceutical-related immune complexes: review and case studies.

PubMed

Rojko, Jennifer L; Evans, Mark G; Price, Shari A; Han, Bora; Waine, Gary; DeWitte, Mark; Haynes, Jill; Freimark, Bruce; Martin, Pauline; Raymond, James T; Evering, Winston; Rebelatto, Marlon C; Schenck, Emanuel; Horvath, Christopher

2014-06-01

Vascular inflammation, infusion reactions, glomerulopathies, and other potentially adverse effects may be observed in laboratory animals, including monkeys, on toxicity studies of therapeutic monoclonal antibodies and recombinant human protein drugs. Histopathologic and immunohistochemical (IHC) evaluation suggests these effects may be mediated by deposition of immune complexes (ICs) containing the drug, endogenous immunoglobulin, and/or complement components in the affected tissues. ICs may be observed in glomerulus, blood vessels, synovium, lung, liver, skin, eye, choroid plexus, or other tissues or bound to neutrophils, monocytes/macrophages, or platelets. IC deposition may activate complement, kinin, and/or coagulation/fibrinolytic pathways and result in a systemic proinflammatory response. IC clearance is biphasic in humans and monkeys (first from plasma to liver and/or spleen, second from liver or spleen). IC deposition/clearance is affected by IC composition, immunomodulation, and/or complement activation. Case studies are presented from toxicity study monkeys or rats and indicate IHC-IC deposition patterns similar to those predicted by experimental studies of IC-mediated reactions to heterologous protein administration to monkeys and other species. The IHC-staining patterns are consistent with findings associated with generalized and localized IC-associated pathology in humans. However, manifestations of immunogenicity in preclinical species are generally not considered predictive to humans. © 2014 by The Author(s).

Mechanisms of Host-Pathogen Protein Complex Formation and Bacterial Immune Evasion of Streptococcus suis Protein Fhb.

PubMed

Li, Xueqin; Liu, Peng; Gan, Shuzhen; Zhang, Chunmao; Zheng, Yuling; Jiang, Yongqiang; Yuan, Yuan

2016-08-12

Streptococcus suis serotype 2 (S. suis 2)-induced sepsis and meningitis are often accompanied by bacteremia. The evasion of polymorphonuclear leukocyte-mediated phagocytic clearance is central to the establishment of bacteremia caused by S. suis 2 and is facilitated by the ability of factor H (FH)-binding protein (Fhb) to bind FH on the bacterial surface, thereby impeding alternative pathway complement activation and phagocytic clearance. Here, C3b/C3d was found to bind to Fhb, along with FH, forming a large immune complex. The formation of this immune complex was mediated by domain II of Fhb via electrostatic and hydrophobic interactions, which, to our knowledge, is a new type of interaction. Interestingly, Fhb was found to be associated with the cell envelope and also present in the culture supernatant, where secreted Fhb inhibited complement activation via interactions with domain II, thereby enhancing antiphagocytic clearance by polymorphonuclear leukocytes. Thus, Fhb is a multifunctional bacterial protein, which binds host complement component C3 as well as FH and interferes with innate immune recognition in a secret protein manner. S. suis 2 therefore appears to have developed a new strategy to combat host innate immunity and enhance survival in host blood. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Mechanisms of Host-Pathogen Protein Complex Formation and Bacterial Immune Evasion of Streptococcus suis Protein Fhb*

PubMed Central

Li, Xueqin; Liu, Peng; Gan, Shuzhen; Zhang, Chunmao; Zheng, Yuling; Jiang, Yongqiang; Yuan, Yuan

2016-01-01

Streptococcus suis serotype 2 (S. suis 2)-induced sepsis and meningitis are often accompanied by bacteremia. The evasion of polymorphonuclear leukocyte-mediated phagocytic clearance is central to the establishment of bacteremia caused by S. suis 2 and is facilitated by the ability of factor H (FH)-binding protein (Fhb) to bind FH on the bacterial surface, thereby impeding alternative pathway complement activation and phagocytic clearance. Here, C3b/C3d was found to bind to Fhb, along with FH, forming a large immune complex. The formation of this immune complex was mediated by domain II of Fhb via electrostatic and hydrophobic interactions, which, to our knowledge, is a new type of interaction. Interestingly, Fhb was found to be associated with the cell envelope and also present in the culture supernatant, where secreted Fhb inhibited complement activation via interactions with domain II, thereby enhancing antiphagocytic clearance by polymorphonuclear leukocytes. Thus, Fhb is a multifunctional bacterial protein, which binds host complement component C3 as well as FH and interferes with innate immune recognition in a secret protein manner. S. suis 2 therefore appears to have developed a new strategy to combat host innate immunity and enhance survival in host blood. PMID:27342778

Autocrine Complement Inhibits IL10-Dependent T-cell-Mediated Antitumor Immunity to Promote Tumor Progression.

PubMed

Wang, Yu; Sun, Sheng-Nan; Liu, Qing; Yu, Yang-Yang; Guo, Jian; Wang, Kun; Xing, Bao-Cai; Zheng, Qing-Feng; Campa, Michael J; Patz, Edward F; Li, Shi-You; He, You-Wen

2016-09-01

In contrast to its inhibitory effects on many cells, IL10 activates CD8(+) tumor-infiltrating lymphocytes (TIL) and enhances their antitumor activity. However, CD8(+) TILs do not routinely express IL10, as autocrine complement C3 inhibits IL10 production through complement receptors C3aR and C5aR. CD8(+) TILs from C3-deficient mice, however, express IL10 and exhibit enhanced effector function. C3-deficient mice are resistant to tumor development in a T-cell- and IL10-dependent manner; human TILs expanded with IL2 plus IL10 increase the killing of primary tumors in vitro compared with IL2-treated TILs. Complement-mediated inhibition of antitumor immunity is independent of the programmed death 1/programmed death ligand 1 (PD-1/PD-L1) immune checkpoint pathway. Our findings suggest that complement receptors C3aR and C5aR expressed on CD8(+) TILs represent a novel class of immune checkpoints that could be targeted for tumor immunotherapy. Moreover, incorporation of IL10 in the expansion of TILs and in gene-engineered T cells for adoptive cell therapy enhances their antitumor efficacy. Our data suggest novel strategies to enhance immunotherapies: a combined blockade of complement signaling by antagonists to C3aR, C5aR, and anti-PD-1 to enhance anti-PD-1 efficacy; a targeted IL10 delivery to CD8(+) TILs using anti-PD-1-IL10 or anti-CTLA4-IL10 fusion proteins; and the addition of IL10 in TIL expansion for adoptive cellular therapy. Cancer Discov; 6(9); 1022-35. ©2016 AACR.See related commentary by Peng et al., p. 953This article is highlighted in the In This Issue feature, p. 932. ©2016 American Association for Cancer Research.

A Novel Quantitative Hemolytic Assay Coupled with Restriction Fragment Length Polymorphisms Analysis Enabled Early Diagnosis of Atypical Hemolytic Uremic Syndrome and Identified Unique Predisposing Mutations in Japan

PubMed Central

Yoshida, Yoko; Miyata, Toshiyuki; Matsumoto, Masanori; Shirotani-Ikejima, Hiroko; Uchida, Yumiko; Ohyama, Yoshifumi; Kokubo, Tetsuro; Fujimura, Yoshihiro

2015-01-01

For thrombotic microangiopathies (TMAs), the diagnosis of atypical hemolytic uremic syndrome (aHUS) is made by ruling out Shiga toxin-producing Escherichia coli (STEC)-associated HUS and ADAMTS13 activity-deficient thrombotic thrombocytopenic purpura (TTP), often using the exclusion criteria for secondary TMAs. Nowadays, assays for ADAMTS13 activity and evaluation for STEC infection can be performed within a few hours. However, a confident diagnosis of aHUS often requires comprehensive gene analysis of the alternative complement activation pathway, which usually takes at least several weeks. However, predisposing genetic abnormalities are only identified in approximately 70% of aHUS. To facilitate the diagnosis of complement-mediated aHUS, we describe a quantitative hemolytic assay using sheep red blood cells (RBCs) and human citrated plasma, spiked with or without a novel inhibitory anti-complement factor H (CFH) monoclonal antibody. Among 45 aHUS patients in Japan, 24% (11/45) had moderate-to-severe (≥50%) hemolysis, whereas the remaining 76% (34/45) patients had mild or no hemolysis (<50%). The former group is largely attributed to CFH-related abnormalities, and the latter group has C3-p.I1157T mutations (16/34), which were identified by restriction fragment length polymorphism (RFLP) analysis. Thus, a quantitative hemolytic assay coupled with RFLP analysis enabled the early diagnosis of complement-mediated aHUS in 60% (27/45) of patients in Japan within a week of presentation. We hypothesize that this novel quantitative hemolytic assay would be more useful in a Caucasian population, who may have a higher proportion of CFH mutations than Japanese patients. PMID:25951460

Complement-mediated neutrophil activation in sepsis- and trauma-related adult respiratory distress syndrome. Clarification with radioaerosol lung scans

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tennenberg, S.D.; Jacobs, M.P.; Solomkin, J.S.

Complement-mediated neutrophil activation (CMNA) has been proposed as an important pathogenic mechanism causing acute microvascular lung injury in the adult respiratory distress syndrome (ARDS). To clarify the relationship between CMNA and evolving lung injury, we studied 26 patients with multiple trauma and sepsis within 24 hours of risk establishment for ARDS. Pulmonary alveolar-capillary permeability (PACP) was quantified as the clearance rate of a particulate radioaerosol. Seventeen patients (65%) had increased PACP (six developed ARDS) while nine (35%) had normal PACP (none developed ARDS; clearance rates of 3.4%/min and 1.5%/min, respectively). These patients, regardless of evidence of early lung injury, hadmore » elevated plasma C3adesArg levels and neutrophil chemotactic desensitization to C5a/C5adesArg. Plasma C3adesArg levels correlated weakly, but significantly, with PACP. Thus, CMNA may be a necessary, but not a sufficient, pathogenic mechanism in the evolution of ARDS.« less

Excessive innate immune response and mutant D222G/N in severe A (H1N1) pandemic influenza.

PubMed

Berdal, Jan-Erik; Mollnes, Tom E; Wæhre, Torgun; Olstad, Ole K; Halvorsen, Bente; Ueland, Thor; Laake, Jon H; Furuseth, May T; Maagaard, Anne; Kjekshus, Harald; Aukrust, Pål; Jonassen, Christine M

2011-10-01

Explore the role of viral factors and immune response in patients with severe pandemic pdmH1N1 illness without significant co-morbidity. Seven patients with pdmH1N1 influenza, bilateral chest X-rays infiltrates, requiring mechanical ventilator support were consecutively recruited. Seven age- and gender-matched healthy individuals served as controls. Four patients were viremic, two with the mutant D222G/N pdmH1N1.Microarray analyses of peripheral blood leukocytes suggested a marked granulocytes activation, but no up-regulation of inflammatory cytokine mRNA. Patients with severe pdmH1NI had a marked systemic complement activation, and in contrast to the lack of cytokine mRNA up-regulation in blood leukocytes, plasma levels of a broad range of inflammatory mediators, including IP-10, and mediators involved in pulmonary remodelling were markedly elevated. Patients with mutant virus had particularly high IP-10 levels, and the most pronounced complement activation. In severe pdmH1N1, viremia was common and the D222G/N mutant was found in half of the viremic patients. Host immune response was characterized by strong activation of the innate immune system, including complement and granulocytes activation, increased serum levels of inflammation and pulmonary remodelling markers, possibly contributing to the observed tissue damage. However, few patients were included and further studies are needed to characterize the immune response in severe pdmH1N1 infection. Copyright © 2011 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

Antimicrobial strategies centered around reactive oxygen species - bactericidal antibiotics, photodynamic therapy and beyond

PubMed Central

Vatansever, Fatma; de Melo, Wanessa C.M.A.; Avci, Pinar; Vecchio, Daniela; Sadasivam, Magesh; Gupta, Asheesh; Chandran, Rakkiyappan; Karimi, Mahdi; Parizotto, Nivaldo A; Yin, Rui; Tegos, George P; Hamblin, Michael R

2013-01-01

Reactive oxygen species (ROS) can attack a diverse range of targets to exert antimicrobial activity, which accounts for their versatility in mediating host defense against a broad range of pathogens. Most ROS are formed by the partial reduction of molecular oxygen. Four major ROS are recognized comprising: superoxide (O2•−), hydrogen peroxide (H2O2), hydroxyl radical (•OH), and singlet oxygen (1O2), but they display very different kinetics and levels of activity. The effects of O2•− and H2O2 are less acute than those of •OH and 1O2, since the former are much less reactive and can be detoxified by endogenous antioxidants (both enzymatic and non-enzymatic) that are induced by oxidative stress. In contrast, no enzyme can detoxify •OH or 1O2, making them extremely toxic and acutely lethal. The present review will highlight the various methods of ROS formation and their mechanism of action. Antioxidant defenses against ROS in microbial cells and the use of ROS by antimicrobial host defense systems are covered. Antimicrobial approaches primarily utilizing ROS comprise both bactericidal antibiotics, and non-pharmacological methods such as photodynamic therapy, titanium dioxide photocatalysis, cold plasma and medicinal honey. A brief final section covers, reactive nitrogen species, and related therapeutics, such as acidified nitrite and nitric oxide releasing nanoparticles. PMID:23802986

Nitric oxide-releasing poly(lactic-co-glycolic acid)-polyethylenimine nanoparticles for prolonged nitric oxide release, antibacterial efficacy, and in vivo wound healing activity

PubMed Central

Nurhasni, Hasan; Cao, Jiafu; Choi, Moonjeong; Kim, Il; Lee, Bok Luel; Jung, Yunjin; Yoo, Jin-Wook

2015-01-01

Nitric oxide (NO)-releasing nanoparticles (NPs) have emerged as a wound healing enhancer and a novel antibacterial agent that can circumvent antibiotic resistance. However, the NO release from NPs over extended periods of time is still inadequate for clinical application. In this study, we developed NO-releasing poly(lactic-co-glycolic acid)-polyethylenimine (PEI) NPs (NO/PPNPs) composed of poly(lactic-co-glycolic acid) and PEI/diazeniumdiolate (PEI/NONOate) for prolonged NO release, antibacterial efficacy, and wound healing activity. Successful preparation of PEI/NONOate was confirmed by proton nuclear magnetic resonance, Fourier transform infrared spectroscopy, and ultraviolet/visible spectrophotometry. NO/PPNPs were characterized by particle size, surface charge, and NO loading. The NO/PPNPs showed a prolonged NO release profile over 6 days without any burst release. The NO/PPNPs exhibited potent bactericidal efficacy against methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa concentration-dependently and showed the ability to bind on the surface of the bacteria. We also found that the NO released from the NO/PPNPs mediates bactericidal efficacy and is not toxic to healthy fibroblast cells. Furthermore, NO/PPNPs accelerated wound healing and epithelialization in a mouse model of a MRSA-infected wound. Therefore, our results suggest that the NO/PPNPs presented in this study could be a suitable approach for treating wounds and various skin infections. PMID:25960648

Oxygenated monoterpenes citral and carvacrol cause oxidative damage in Escherichia coli without the involvement of tricarboxylic acid cycle and Fenton reaction.

PubMed

Chueca, Beatriz; Pagán, Rafael; García-Gonzalo, Diego

2014-10-17

Oxygenated monoterpenes citral and carvacrol are common constituents of many essential oils (EOs) that have been extensively studied as antimicrobial agents but whose mechanisms of microbial inactivation have not been totally elucidated. A recent study described a mechanism of Escherichia coli death for (+)-limonene, a hydrocarbon monoterpene also frequently present in EOs, similar to the common mechanism proposed for bactericidal antibiotics. This mechanism involves the formation of Fenton-mediated hydroxyl radical, a reactive oxygen species (ROS), via tricarboxylic acid (TCA) cycle, which would ultimately inactivate cells. Our objective was to determine whether E. coli MG1655 inactivation by citral and carvacrol follows a similar mechanism of cell death. Challenging experiments with 300μL/L citral and 100μL/L carvacrol inactivated at least 2.5log10cycles of exponentially growing cells in 3h under aerobic conditions. The presence of thiourea (an ROS scavenger) reduced cell inactivation in 2log10cycles, demonstrating the role of ROS in cell death. Decreased resistance of a ΔrecA mutant (deficient in an enzyme involved in SOS response to DNA damage) indicated that citral and carvacrol caused oxidative damage to DNA. Although the mechanism of E. coli inactivation by carvacrol and citral was similarly mediated by ROS, their formation did not follow the same pathways described for (+)-limonene and bactericidal drugs because neither Fenton reaction nor NADH production via the TCA cycle was involved in cell death. Moreover, further experiments demonstrated antimicrobial activity of citral and carvacrol in anaerobic environments without the involvement of ROS. As a consequence, cell death by carvacrol and citral in anaerobiosis follows a different mechanism than that observed under aerobic conditions. These results demonstrated a different mechanism of inactivation by citral and carvacrol with regard to (+)-limonene and bactericidal antibiotics, indicating the complexity of the mechanisms of bacterial inactivation among EO constituents. Advancements in the description of these mechanisms will help in extending and improving the use of these compounds as natural antimicrobials. Copyright © 2014 Elsevier B.V. All rights reserved.

Enhanced susceptibility to acute pneumococcal otitis media in mice deficient in complement C1qa, factor B, and factor B/C2.

PubMed

Tong, Hua Hua; Li, Yong Xing; Stahl, Gregory L; Thurman, Joshua M

2010-03-01

To define the roles of specific complement activation pathways in host defense against Streptococcus pneumoniae in acute otitis media (AOM), we investigated the susceptibility to AOM in mice deficient in complement factor B and C2 (Bf/C2(-/)(-)), C1qa (C1qa(-/)(-)), and factor B (Bf(-)(/)(-)). Bacterial titers of both S. pneumoniae serotype 6A and 14 in the middle ear lavage fluid samples from Bf/C2(-/)(-), Bf(-)(/)(-), and C1qa(-/)(-) mice were significantly higher than in samples from wild-type mice 24 h after transtympanical infection (P < 0.05) and remained persistently higher in samples from Bf/C2(-/)(-) mice than in samples from wild-type mice. Bacteremia occurred in Bf/C2(-/)(-), Bf(-)(/)(-), and C1qa(-/)(-) mice infected with both strains, but not in wild-type mice. Recruitment of inflammatory cells was paralleled by enhanced production of inflammatory mediators in the middle ear lavage samples from Bf/C2(-/)(-) mice. C3b deposition on both strains was greatest for sera obtained from wild-type mice, followed by C1qa(-)(/)(-) and Bf(-)(/)(-) mice, and least for Bf/C2(-)(/)(-) mice. Opsonophagocytosis and whole-blood killing capacity of both strains were significantly decreased in the presence of sera or whole blood from complement-deficient mice compared to wild-type mice. These findings indicate that both the classical and alternative complement pathways are critical for middle ear immune defense against S. pneumoniae. The reduced capacity of complement-mediated opsonization and phagocytosis in the complement-deficient mice appears to be responsible for the impaired clearance of S. pneumoniae from the middle ear and dissemination to the bloodstream during AOM.

Construction and characterization of an anti-CD20 mAb nanocomb with exceptionally excellent lymphoma-suppressing activity.

PubMed

Li, Hua-Fei; Wu, Cong; Chen, Ting; Zhang, Ge; Zhao, He; Ke, Chang-Hong; Xu, Zheng

2015-01-01

The CD20-directed monoclonal antibody rituximab (RTX) established a new era in the treatment of non-Hodgkin lymphoma (NHL); however, suboptimal response and/or resistance to RTX still limit its clinical merits. Although four effector mechanisms are validated to participate in CD20-based immunotherapy, including complement-dependent cytotoxicity, antibody-dependent cell-mediated cytotoxicity, caspase-dependent apoptosis, and lysosome-mediated programmed cell death (PCD), they could hardly be synchronously activated by any anti-CD20 mAb or mAb derivative until now. Herein, a novel mAb nanocomb (polyethylenimine polymer-RTX-tositumomab [PPRT nanocomb]) was firstly constructed through mass arming two different anti-CD20 mAbs (RTX and tositumomab) to one polymer by nanotechnology. Comparing with free mAbs, PPRT nanocomb possesses a comparable binding ability and reduced "off-rate" to surface CD20 of NHL cells. When treated by PPRT nanocomb, the caspase-dependent apoptosis was remarkably enhanced except for concurrently eliciting complement-dependent cytotoxicity, antibody-dependent cell-mediated cytotoxicity, and lysosome-mediated PCD. Besides, "cross-cell link"-assisted homotypic adhesion by PPRT nanocomb further enhanced the susceptibility to PCD of lymphoma cells. Pharmacokinetic assays revealed that PPRT nanocomb experienced a relatively reduced clearance from peripheral blood compared with free antibodies. With the cooperation of all the abovementioned superiorities, PPRT nanocomb exhibits exceptionally excellent in vivo antitumor activities in both disseminated and localized human NHL xenotransplant models.

Construction and characterization of an anti-CD20 mAb nanocomb with exceptionally excellent lymphoma-suppressing activity

PubMed Central

Li, Hua-Fei; Wu, Cong; Chen, Ting; Zhang, Ge; Zhao, He; Ke, Chang-Hong; Xu, Zheng

2015-01-01

The CD20-directed monoclonal antibody rituximab (RTX) established a new era in the treatment of non-Hodgkin lymphoma (NHL); however, suboptimal response and/or resistance to RTX still limit its clinical merits. Although four effector mechanisms are validated to participate in CD20-based immunotherapy, including complement-dependent cytotoxicity, antibody-dependent cell-mediated cytotoxicity, caspase-dependent apoptosis, and lysosome-mediated programmed cell death (PCD), they could hardly be synchronously activated by any anti-CD20 mAb or mAb derivative until now. Herein, a novel mAb nanocomb (polyethylenimine polymer–RTX–tositumomab [PPRT nanocomb]) was firstly constructed through mass arming two different anti-CD20 mAbs (RTX and tositumomab) to one polymer by nanotechnology. Comparing with free mAbs, PPRT nanocomb possesses a comparable binding ability and reduced “off-rate” to surface CD20 of NHL cells. When treated by PPRT nanocomb, the caspase-dependent apoptosis was remarkably enhanced except for concurrently eliciting complement-dependent cytotoxicity, antibody-dependent cell-mediated cytotoxicity, and lysosome-mediated PCD. Besides, “cross-cell link”-assisted homotypic adhesion by PPRT nanocomb further enhanced the susceptibility to PCD of lymphoma cells. Pharmacokinetic assays revealed that PPRT nanocomb experienced a relatively reduced clearance from peripheral blood compared with free antibodies. With the cooperation of all the abovementioned superiorities, PPRT nanocomb exhibits exceptionally excellent in vivo antitumor activities in both disseminated and localized human NHL xenotransplant models. PMID:26257518

C1q-targeted inhibition of the classical complement pathway prevents injury in a novel mouse model of acute motor axonal neuropathy.

PubMed

McGonigal, Rhona; Cunningham, Madeleine E; Yao, Denggao; Barrie, Jennifer A; Sankaranarayanan, Sethu; Fewou, Simon N; Furukawa, Koichi; Yednock, Ted A; Willison, Hugh J

2016-03-02

Guillain-Barré syndrome (GBS) is an autoimmune disease that results in acute paralysis through inflammatory attack on peripheral nerves, and currently has limited, non-specific treatment options. The pathogenesis of the acute motor axonal neuropathy (AMAN) variant is mediated by complement-fixing anti-ganglioside antibodies that directly bind and injure the axon at sites of vulnerability such as nodes of Ranvier and nerve terminals. Consequently, the complement cascade is an attractive target to reduce disease severity. Recently, C5 complement component inhibitors that block the formation of the membrane attack complex and subsequent downstream injury have been shown to be efficacious in an in vivo anti-GQ1b antibody-mediated mouse model of the GBS variant Miller Fisher syndrome (MFS). However, since gangliosides are widely expressed in neurons and glial cells, injury in this model was not targeted exclusively to the axon and there are currently no pure mouse models for AMAN. Additionally, C5 inhibition does not prevent the production of early complement fragments such as C3a and C3b that can be deleterious via their known role in immune cell and macrophage recruitment to sites of neuronal damage. In this study, we first developed a new in vivo transgenic mouse model of AMAN using mice that express complex gangliosides exclusively in neurons, thereby enabling specific targeting of axons with anti-ganglioside antibodies. Secondly, we have evaluated the efficacy of a novel anti-C1q antibody (M1) that blocks initiation of the classical complement cascade, in both the newly developed anti-GM1 antibody-mediated AMAN model and our established MFS model in vivo. Anti-C1q monoclonal antibody treatment attenuated complement cascade activation and deposition, reduced immune cell recruitment and axonal injury, in both mouse models of GBS, along with improvement in respiratory function. These results demonstrate that neutralising C1q function attenuates injury with a consequent neuroprotective effect in acute GBS models and promises to be a useful new target for human therapy.

EFFECT OF INORGANIC CATIONS ON BACTERICIDAL ACTIVITY OF ANIONIC SURFACTANTS

PubMed Central

Voss, J. G.

1963-01-01

Voss, J. G. (Procter & Gamble Co., Cincinnati, Ohio). Effect of inorganic cations on bactericidal activity of anionic surfactants. J. Bacteriol. 86:207–211. 1963.—The bactericidal effectiveness of two alkyl benzene sulfonates and of three other types of anionic surfactants against Staphylococcus aureus is increased in the presence of low concentrations of divalent cations, especially alkaline earths and metals of group IIB of the periodic table. The cations may act by decreasing the negative charge at the cell surface and increasing adsorption of the surfactant anions, leading to damage to the cytoplasmic membrane and death of the cell. Increased adsorption of surfactant is also found with Escherichia coli, but does not lead to death of the cell. PMID:14058942

Synthesis and characterization of bactericidal silver nanoparticles using cultural filtrate of simulated microgravity grown Klebsiella pneumoniae.

PubMed

Kalpana, Duraisamy; Lee, Yang Soo

2013-03-05

Silver nanoparticles were synthesized by biological method using cultural filtrate of Klebsiella pneumoniae cultured under simulated microgravity and silver nitrate solution as precursor. The nanoparticles exhibited typical plasmon absorption maximum of silver nanoparticles between 405 and 407 nm. Spherical silver nanoparticles were found to have size between 15 and 37 nm by TEM analysis. XRD pattern corresponding to planes (111), (200), (220) (311) revealed the crystalline nature of the biosynthesized silver nanoparticles. FTIR spectrum proposed stabilization of silver nanoparticles by the protein molecules present in the cultural filtrate. The silver nanoparticles exhibited high bactericidal activity against Salmonella enterica, Escherichia coli and moderate bactericidal activity against Streptococcus pyogenes. Copyright © 2012 Elsevier Inc. All rights reserved.

Complement Activation in Relation to Capillary Leakage in Children with Septic Shock and Purpura

PubMed Central

Hazelzet, Jan A.; de Groot, Ronald; van Mierlo, Gerard; Joosten, Koen F. M.; van der Voort, Edwin; Eerenberg, Anke; Suur, Marja H.; Hop, Wim C. J.; Hack, C. Erik

1998-01-01

To assess the relationship between capillary leakage and inflammatory mediators during sepsis, blood samples were taken on hospital admission, as well as 24 and 72 h later, from 52 children (median age, 3.3 years) with severe meningococcal sepsis, of whom 38 survived and 14 died. Parameters related to cytokines (interleukin 6 [IL-6] IL-8, plasma phospholipase A2, and C-reactive protein [CRP]), to neutrophil degranulation (elastase and lactoferrin), to complement activation (C3a, C3b/c, C4b/c, and C3- and C4-CRP complexes), and to complement regulation (functional and inactivated C1 inhibitor and C4BP) were determined. The degree of capillary leakage was derived from the amount of plasma infused and the severity of disease by assessing the pediatric risk of mortality (PRISM) score. Levels of IL-6, IL-8, C3b/c, C3-CRP complexes, and C4BP on admission, adjusted for the duration of skin lesions, were significantly different in survivors and nonsurvivors (C3b/c levels were on average 2.2 times higher in nonsurvivors, and C3-CRP levels were 1.9 times higher in survivors). Mortality was independently related to the levels of C3b/c and C3-CRP complexes. In agreement with this, levels of complement activation products correlated well with the PRISM score or capillary leakage. Thus, these data show that complement activation in patients with severe meningococcal sepsis is associated with a poor outcome and a more severe disease course. Further studies should reveal whether complement activation may be a target for therapeutical intervention in this disease. PMID:9784543

Antimicrobial activity of wild mushroom extracts against clinical isolates resistant to different antibiotics.

PubMed

Alves, M J; Ferreira, I C F R; Martins, A; Pintado, M

2012-08-01

This work aimed to screen the antimicrobial activity of aqueous methanolic extracts of 13 mushroom species, collected in Bragança, against several clinical isolates obtained in Hospital Center of Trás-os-Montes and Alto Douro, Portugal. Microdilution method was used to determine the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC). MIC results showed that Russula delica and Fistulina hepatica extracts inhibited the growth of gram-negative (Escherichia coli, Morganella morganni and Pasteurella multocida) and gram-positive (Staphylococcus aureus, MRSA, Enterococcus faecalis, Listeria monocytogenes, Streptococcus agalactiae and Streptococcus pyogenes) bacteria. A bactericide effect of both extracts was observed in Past. multocida, Strep. agalactiae and Strep. pyogenes with MBC of 20, 10 and 5 mg ml⁻¹, respectively. Lepista nuda extract exhibited a bactericide effect upon Past. multocida at 5 mg ml⁻¹ and inhibited Proteus mirabilis at 20 mg ml⁻¹. Ramaria botrytis extract showed activity against Enterococcus faecalis and L. monocytogenes, being bactericide for Past. multocida, Strep. agalactiae (MBCs 20 mg ml⁻¹) and Strep. pyogenes (MBC 10 mg ml⁻¹). Leucopaxillus giganteus extract inhibited the growth of E. coli and Pr. mirabilis, being bactericide for Past. multocida, Strep. pyogenes and Strep. agalactiae. Fistulina hepatica, R. botrytis and R. delica are the most promising species as antimicrobial agents. Mushroom extracts could be an alternative as antimicrobials against pathogenic micro-organisms resistant to conventional treatments. © 2012The Authors Journal of Applied Microbiology © 2012 The Society for Applied Microbiology.

Urease from Helicobacter pylori is inactivated by sulforaphane and other isothiocyanates

PubMed Central

Fahey, Jed W.; Stephenson, Katherine K.; Wade, Kristina L.; Talalay, Paul

2013-01-01

Infections by Helicobacter pylori are very common, causing gastroduodenal inflammation including peptic ulcers, and increasing the risk of gastric neoplasia. The isothiocyanate (ITC) sulforaphane [SF; 1-isothiocyanato-4-(methylsulfinyl)butane] derived from edible crucifers such as broccoli is potently bactericidal against Helicobacter, including antibiotic-resistant strains, suggesting a possible dietary therapy. Gastric H. pylori infections express high urease activity which generates ammonia, neutralizes gastric acidity, and promotes inflammation. The finding that SF inhibits (inactivates) urease (jack bean and Helicobacter) raised the issue of whether these properties might be functionally related. The rates of inactivation of urease activity depend on enzyme and SF concentrations and show first order kinetics. Treatment with SF results in time-dependent increases in the ultraviolet absorption of partially purified Helicobacter urease in the 280–340 nm region. This provides direct spectroscopic evidence for the formation of dithiocarbamates between the ITC group of SF and cysteine thiols of urease. The potencies of inactivation of Helicobacter urease by isothiocyanates structurally related to SF were surprisingly variable. Natural isothiocyanates closely related to SF, previously shown to be bactericidal (berteroin, hirsutin, phenethyl isothiocyanate, alyssin, and erucin), did not inactivate urease activity. Furthermore, SF is bactericidal against both urease positive and negative H. pylori strains. In contrast, some isothiocyanates such as benzoyl-ITC, are very potent urease inactivators, but are not bactericidal. The bactericidal effects of SF and other ITC against Helicobacter are therefore not obligatorily linked to urease inactivation, but may reduce the inflammatory component of Helicobacter infections. PMID:23583386

Availability of the basal planes of graphene oxide determines whether it is antibacterial.

PubMed

Hui, Liwei; Piao, Ji-Gang; Auletta, Jeffrey; Hu, Kan; Zhu, Yanwu; Meyer, Tara; Liu, Haitao; Yang, Lihua

2014-08-13

There are significant controversies on the antibacterial properties of graphene oxide (GO): GO was reported to be bactericidal in saline, whereas its activity in nutrient broth was controversial. To unveil the mechanisms underlying these contradictions, we performed antibacterial assays under comparable conditions. In saline, bare GO sheets were intrinsically bactericidal, yielding a bacterial survival percentage of <1% at 200 μg/mL. Supplementing saline with ≤10% Luria-Bertani (LB) broth, however, progressively deactivated its bactericidal activity depending on LB-supplementation ratio. Supplementation of 10% LB made GO completely inactive; instead, ∼100-fold bacterial growth was observed. Atomic force microscopy images showed that certain LB components were adsorbed on GO basal planes. Using bovine serum albumin and tryptophan as well-defined model adsorbates, we found that noncovalent adsorption on GO basal planes may account for the deactivation of GO's bactericidal activity. Moreover, this deactivation mechanism was shown to be extrapolatable to GO's cytotoxicity against mammalian cells. Taken together, our observations suggest that bare GO intrinsically kills both bacteria and mammalian cells and noncovalent adsorption on its basal planes may be a global deactivation mechanism for GO's cytotoxicity.

Naturally occurring anti-Salmonella agents.

PubMed

Kubo, I; Fujita, K

2001-12-01

Polygodial and (2E)-hexenal were found to possess antibacterial activity against Salmonella choleraesuis with the minimum bactericidal concentrations (MBC) of 50 microg/mL (0.17 mM) and 100 microg/mL (0.98 mM), respectively. The time kill curve study showed that these two alpha,beta-unsaturated aldehydes were bactericidal against this food-borne bacterium at any stage of growth. However, they showed different effects on the growth of S. choleraesuis. The combination of polygodial and anethole exhibited strong synergism on their bacteriostatic action but only marginal synergism on their bactericidal action.

Complement Protein C1q Modulates Neurite Outgrowth In Vitro and Spinal Cord Axon Regeneration In Vivo

PubMed Central

Peterson, Sheri L.; Nguyen, Hal X.; Mendez, Oscar A.

2015-01-01

Traumatic injury to CNS fiber tracts is accompanied by failure of severed axons to regenerate and results in lifelong functional deficits. The inflammatory response to CNS trauma is mediated by a diverse set of cells and proteins with varied, overlapping, and opposing effects on histological and behavioral recovery. Importantly, the contribution of individual inflammatory complement proteins to spinal cord injury (SCI) pathology is not well understood. Although the presence of complement components increases after SCI in association with axons and myelin, it is unknown whether complement proteins affect axon growth or regeneration. We report a novel role for complement C1q in neurite outgrowth in vitro and axon regrowth after SCI. In culture, C1q increased neurite length on myelin. Protein and molecular assays revealed that C1q interacts directly with myelin associated glycoprotein (MAG) in myelin, resulting in reduced activation of growth inhibitory signaling in neurons. In agreement with a C1q-outgrowth-enhancing mechanism in which C1q binding to MAG reduces MAG signaling to neurons, complement C1q blocked both the growth inhibitory and repulsive turning effects of MAG in vitro. Furthermore, C1q KO mice demonstrated increased sensory axon turning within the spinal cord lesion after SCI with peripheral conditioning injury, consistent with C1q-mediated neutralization of MAG. Finally, we present data that extend the role for C1q in axon growth and guidance to include the sprouting patterns of descending corticospinal tract axons into spinal gray matter after dorsal column transection SCI. PMID:25762679

Bactericidal activity of herbal volatile oil extracts against multidrug-resistant Acinetobacter baumannii

PubMed Central

Intorasoot, Amornrat; Chornchoem, Piyaorn; Sookkhee, Siriwoot; Intorasoot, Sorasak

2017-01-01

Aim: The aim of the study is to investigate the antibacterial activity of 10 volatile oils extracted from medicinal plants, including galangal (Alpinia galanga Linn.), ginger (Zingiber officinale), plai (Zingiber cassumunar Roxb.), lime (Citrus aurantifolia), kaffir lime (Citrus hystrix DC.), sweet basil (Ocimum basilicum Linn.), tree basil (Ocimum gratissimum), lemongrass (Cymbopogon citratus DC.), clove (Syzygium aromaticum), and cinnamon (Cinnamomum verum) against four standard strains of Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Acinetobacter baumannii, and 30 clinical isolates of multidrug-resistant A. baumannii (MDR-A. baumannii). Materials and Methods: Agar diffusion, minimum inhibitory concentration, and minimum bactericidal concentration (MBC) were employed for the determination of bactericidal activity of water distilled medicinal plants. Tea tree oil (Melaleuca alternifolia) was used as positive control in this study. Results: The results indicated the volatile oil extracted from cinnamon exhibited potent antibacterial activity against the most common human pathogens, S. aureus, E. coli, P. aeruginosa, and A. baumannii. Most of volatile oil extracts were less effective against non-fermentative bacteria, P. aeruginosa. In addition, volatile oil extracted from cinnamon, clove, and tree basil possessed potent bactericidal activity against MDR-A. baumannii with MBC90 of 0.5, 1, and 2 mg/mL, respectively. Conclusions: The volatile oil extracts would be useful as alternative natural product for the treatment of the most common human pathogens and MDR-A. baumannii infections. PMID:28512603

Neutralization of bactericidal activity related to antimicrobial carry-over in broiler carcass rinse samples

USDA-ARS?s Scientific Manuscript database

Chemical antimicrobial interventions used as poultry processing aids may have potential for carry-over into whole poultry carcass buffered peptone water (BPW) rinses collected for the recovery of viable Salmonella. Such carry-over may lead to false negative indications due to continuing bactericidal...

Antagonistic effects of lipids against the bactericidal activity of thymol-beta-D-glucopyranoside

USDA-ARS?s Scientific Manuscript database

The gut of food-producing animals is a reservoir for zoonotic pathogens. Thymol is bactericidal against Salmonella, E. coli, and Campylobacter, but its rapid absorption from the proximal gut reveals a need for protective technologies to deliver effective concentrations to the lower gut where the pa...

Complement C3 and C5 play critical roles in traumatic brain cryoinjury: blocking effects on neutrophil extravasation by C5a receptor antagonist☆

PubMed Central

Sewell, Diane L.; Nacewicz, Brendon; Liu, Frances; Macvilay, Sinarack; Erdei, Anna; Lambris, John D.; Sandor, Matyas; Fabry, Zsuzsa

2016-01-01

The role of complement components in traumatic brain injury is poorly understood. Here we show that secondary damage after acute cryoinjury is significantly reduced in C3−/− or C5−/− mice or in mice treated with C5a receptor antagonist peptides. Injury sizes and neutrophil extravasation were compared. While neutrophil density increased following traumatic brain injury in wild type (C57BL/6) mice, C3-deficient mice demonstrated lower neutrophil extravasation and injury sizes in the brain. RNase protection assay indicated that C3 contributes to the induction of brain inflammatory mediators, MIF, RANTES (CCL5) and MCP-1 (CCL2). Intracranial C3 injection induced neutrophil extravasation in injured brains of C3−/− mice suggesting locally produced C3 is important in brain inflammation. We show that neutrophil extravasation is significantly reduced in both C5−/− mice and C5a receptor antagonist treated cryoinjured mice suggesting that one of the possible mechanisms of C3 effect on neutrophil extravasation is mediated via downstream complement activation products such as C5a. Our data indicates that complement inhibitors may ameliorate traumatic brain injury. PMID:15342196

Antibacterial activity of vegetables and juices.

PubMed

Lee, Yee-Lean; Cesario, Thomas; Wang, Yang; Shanbrom, Edward; Thrupp, Lauri

2003-01-01

We evaluated the antibacterial activities of various fruit and vegetable extracts on common potential pathogens including antibiotic-resistant strains. Standardized bacterial inocula were added to serial dilutions of sterile vegetable and fruit extracts in broth, with final bacterial concentrations of 10(4-5) cells/mL. After overnight incubation at 35 degrees C, antibacterial activity was measured by minimum inhibitory and minimum bactericidal dilutions (for raw juices) or concentrations (for tea). Among the vegetable and fruit extracts tested, all green vegetables showed no antibacterial activity on Staphylococcus epidermidis and Klebsiella pneumoniae. All purple and red vegetable and fruit juices had antibacterial activities in dilutions ranging from 1:2 to 1:16. Garlic juice had significant activity, with bactericidal action in dilutions ranging up to 1:128 of the original juice. Tea also had significant activity, with bactericidal action in concentrations ranging up to 1.6 mg/mL, against a spectrum of pathogens including resistant strains such as methicillin- and ciprofloxacin-resistant staphylococci, vancomycin-resistant enterococci, and ciprofloxacin-resistant Pseudomonas aeruginosa. Tea and garlic have the potential for exploration of broader applications as antibacterial agents.

Arachidonic acid, but not its metabolites, is essential for FcγR-stimulated intracellular killing of Staphylococcus aureus by human monocytes

PubMed Central

Zheng, L; Zomerdijk, T P L; Van Den Barselaar, M T; Geertsma, M F; Van Furth, R; Nibbering, P H

1999-01-01

Since arachidonic acid (AA) production by phospholipase A2 (PLA2) is essential for the Fcγ receptor (FcγR)-mediated respiratory burst and phagocytosis of opsonized erythrocytes by monocytes and macrophages, we focused in this study on the role of AA and its metabolites in the FcγR-stimulated intracellular killing of Staphylococcus aureus by human monocytes. The results revealed that the PLA2 inhibitors, but not inhibitors of cyclo-oxygenase and lipoxygenase, markedly suppressed the FcγR-mediated killing process. The production of O−2 by monocytes upon FcγR cross-linking was inhibited by 4-bromophenacyl bromide in a dose-dependent fashion, indicating that inhibition of PLA2 activity impairs the oxygen-dependent bactericidal mechanisms of monocytes, which could be partially restored by addition of exogenous AA and docosahexaenoic acid, but not myristic acid. These polyunsaturated fatty acids, but not myristic acid, stimulated the intracellular killing of S. aureus by monocytes, although not as effectively as FcγR cross-linking. Furthermore, FcγR cross-linking stimulated the release of AA from monocytes. Studies with selective inhibitors revealed that the FcγR-mediated activation of PLA2 is dependent on Ca2+ and tyrosine kinase activity. Together these results indicate a key role for PLA2/AA, but not its major metabolites, in mediating the FcγR-stimulated intracellular killing of S. aureus by monocytes. PMID:10233682

Influence of nanoscale topology on bactericidal efficiency of black silicon surfaces

NASA Astrophysics Data System (ADS)

Linklater, Denver P.; Khuong Duy Nguyen, Huu; Bhadra, Chris M.; Juodkazis, Saulius; Ivanova, Elena P.

2017-06-01

The nanostructuring of materials to create bactericidal and antibiofouling surfaces presents an exciting alternative to common methods of preventing bacterial adhesion. The fabrication of synthetic bactericidal surfaces has been inspired by the anti-wetting and anti-biofouling properties of insect wings, and other topologies found in nature. Black silicon is one such synthetic surfaces which has established bactericidal properties. In this study we show that time-dependent plasma etching of silicon wafers using 15, 30, and 45 min etching intervals, is able to produce different surface geometries with linearly increasing heights of approximately 280, 430, and 610 nm, respectively. After incubation on these surfaces with Gram-positive Staphylococcus aureus and Gram-negative Pseudomonas aeruginosa bacterial cells it was established that smaller, more densely packed pillars exhibited the greatest bactericidal activity with 85% and 89% inactivation of bacterial cells, respectively. The decrease in the pillar heights, pillar cap diameter and inter-pillar spacing corresponded to a subsequent decrease in the number of attached cells for both bacterial species.

Bactericidal activity of self-assembled palmitic and stearic fatty acid crystals on highly ordered pyrolytic graphite.

PubMed

Ivanova, Elena P; Nguyen, Song Ha; Guo, Yachong; Baulin, Vladimir A; Webb, Hayden K; Truong, Vi Khanh; Wandiyanto, Jason V; Garvey, Christopher J; Mahon, Peter J; Mainwaring, David E; Crawford, Russell J

2017-09-01

The wings of insects such as cicadas and dragonflies have been found to possess nanostructure arrays that are assembled from fatty acids. These arrays can physically interact with the bacterial cell membranes, leading to the death of the cell. Such mechanobactericidal surfaces are of significant interest, as they can kill bacteria without the need for antibacterial chemicals. Here, we report on the bactericidal effect of two of the main lipid components of the insect wing epicuticle, palmitic (C16) and stearic (C18) fatty acids. Films of these fatty acids were re-crystallised on the surface of highly ordered pyrolytic graphite. It appeared that the presence of two additional CH 2 groups in the alkyl chain resulted in the formation of different surface structures. Scanning electron microscopy and atomic force microscopy showed that the palmitic acid microcrystallites were more asymmetric than those of the stearic acid, where the palmitic acid microcrystallites were observed to be an angular abutment in the scanning electron micrographs. The principal differences between the two types of long-chain saturated fatty acid crystallites were the larger density of peaks in the upper contact plane of the palmitic acid crystallites, as well as their greater proportion of asymmetrical shapes, in comparison to that of the stearic acid film. These two parameters might contribute to higher bactericidal activity on surfaces derived from palmitic acid. Both the palmitic and stearic acid crystallite surfaces displayed activity against Gram-negative, rod-shaped Pseudomonas aeruginosa and Gram-positive, spherical Staphylococcus aureus cells. These microcrystallite interfaces might be a useful tool in the fabrication of effective bactericidal nanocoatings. Nanostructured cicada and dragonfly wing surfaces have been discovered to be able physically kill bacterial cells. Here, we report on the successful fabrication of bactericidal three-dimensional structures of two main lipid components of the epicuticle of insect wings, palmitic (C16) and stearic (C18) acids. After crystallisation onto highly ordered pyrolytic graphite, both the palmitic and stearic acid films displayed bactericidal activity against both Gram-negative Pseudomonas aeruginosa and Gram-positive Staphylococcus aureus cells. The simplicity of the production of these microcrystallite interfaces suggests that a fabrication technique, based on solution deposition, could be an effective technique for the application of bactericidal nanocoatings. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

[In vitro antibacterial activity of a new parenteral penem, sulopenem].

PubMed

Yoshida, T; Tateda, E; Hiramatsu, K; Yokota, T

1996-04-01

Eighty percent minimum inhibitory concentrations (MIC80) of sulopenem against clinically isolated 12 to 80 strains of each of different bacteria were as follows: methicillin-susceptible Staphylococcus aureus (MSSA): 0.20 micrograms/ml, methicillin-resistant S. aureus (MRSA): 50 micrograms/ml, coagulase-negative staphylococci: 3.13 micrograms/ml, Streptococcus pyogenes: < or = 0.013 microgram/ml, Streptococcus pneumoniae: < or = 0.013 microgram/ml, beta-streptococci: 0.05 microgram/ml, Enterococcus faecalis: 12.5 micrograms/ml, Enterococcus faecium: > 100 micrograms/ml, Escherichia coli CS2(R+): 0.10 microgram/ml, Klebsiella pneumoniae: 0.05 microgram/ml, Proteus mirabilis: 0.10 microgram/ml, Proteus vulgaris: 0.20 microgram/ml, Morganella morganii: 0.39 micrograms/ml, Providencia rettgeri: 3.13 micrograms/ml, Citrobacter freundii: 0.20 microgram/ml, Enterobacter cloacae: 0.39 microgram/ml, Serratia marcescens: 1.56 micrograms/ml, Pseudomonas aeruginosa: 50 micrograms/ml, Pseudomonas cepacia: 3.13 micrograms/ml, Xanthomonas maltophilia: > 100 micrograms/ml, Acinetobacter calcoaceticus: 1.56 micrograms/ml, ampicillin-resistant Haemophilus influenzae: 0.39 microgram/ml and Bacteroides fragil is: 0.20 microgram/ml, respectively. Sulopenem possesses a stronger activity than flomoxef or cefuzonam against Gram-positive bacteria, the strongest activity among the antibiotics tested against Gram-negative bacteria except P. aeruginosa. Sulopenem has stronger affinities than imipenem to all fractions of PBPs of S. aureus, E. coli, P. vulgaris, S. marcescens, even of P. aeruginosa. Affinities of sulopenem to PBPs-1 and -3 of S. aureus, PBP-2 of E. coli were much stronger than those of imipenem (IPM). Sulopenem generally has small Ki values to all types of beta-lactamases and also has stronger permanent inactivation effect to Ia and IIb types of beta-lactamases than IPM. No synergistic bactericidal activity of sulopenem was apparent with serum complement. However, synergism of sulopenem with macrophages was prominent in bactericidal activity. The cells of E. coli were well phagocytosed and rapidly digested by cultured macrophages in the presence of a higher than 1/8 MIC of sulopenem. Moreover, sulopenem was more stable than imipenem against swine and human dehydropeptidase-Is. Sulopenem is one of the antibiotics that do not induce the appearance of subclones resistant to the drug.

Following the mechanisms of bacteriostatic versus bactericidal action using Raman spectroscopy.

PubMed

Bernatová, Silvie; Samek, Ota; Pilát, Zdeněk; Serý, Mojmír; Ježek, Jan; Jákl, Petr; Siler, Martin; Krzyžánek, Vladislav; Zemánek, Pavel; Holá, Veronika; Dvořáčková, Milada; Růžička, Filip

2013-10-24

Antibiotics cure infections by influencing bacterial growth or viability. Antibiotics can be divided to two groups on the basis of their effect on microbial cells through two main mechanisms, which are either bactericidal or bacteriostatic. Bactericidal antibiotics kill the bacteria and bacteriostatic antibiotics suppress the growth of bacteria (keep them in the stationary phase of growth). One of many factors to predict a favorable clinical outcome of the potential action of antimicrobial chemicals may be provided using in vitro bactericidal/bacteriostatic data (e.g., minimum inhibitory concentrations-MICs). Consequently, MICs are used in clinical situations mainly to confirm resistance, and to determine the in vitro activities of new antimicrobials. We report on the combination of data obtained from MICs with information on microorganisms' "fingerprint" (e.g., DNA/RNA, and proteins) provided by Raman spectroscopy. Thus, we could follow mechanisms of the bacteriostatic versus bactericidal action simply by detecting the Raman bands corresponding to DNA. The Raman spectra of Staphylococcus epidermidis treated with clindamycin (a bacteriostatic agent) indeed show little effect on DNA which is in contrast with the action of ciprofloxacin (a bactericidal agent), where the Raman spectra show a decrease in strength of the signal assigned to DNA, suggesting DNA fragmentation.

Bactericidal activity of culture fluid components of Lactobacillus fermentum strain 90 TS-4 (21) clone 3, and their capacity to modulate adhesion of Candida albicans yeast-like fungi to vaginal epithelial cells.

PubMed

Anokhina, I V; Kravtsov, E G; Protsenko, A V; Yashina, N V; Yermolaev, A V; Chesnokova, V L; Dalin, M V

2007-03-01

Antagonistic activities of L. fermentum strain 90 TS-4 (21), L. casei ATCC 27216, and L. acidophilus ATCC 4356 and bactericidal activity of lactobacillus culture fluid towards E. coli strain K12, S. aureus, and S. epidermidis test cultures were studied. The bactericidal effect of L. fermentum strain 90 TS-4 (21) clone 3 culture fluid preparation (pH 6.0) on the test cultures was dose-dependent. Adhesion of C. albicans yeast-like fungi to vaginal epitheliocytes was more pronounced for strains isolated from women with asymptomatic infection than for strains isolated from women with manifest forms. L. fermentum strain 90 TS-4 (21) clone 3 culture fluid preparation modulated adhesion of yeast-like fungi only if the fungal strain was initially highly adherent.

Targeted complement inhibition as a promising strategy for preventing inflammatory complications in hemodialysis

PubMed Central

DeAngelis, Robert A.; Reis, Edimara S.; Ricklin, Daniel; Lambris, John D.

2012-01-01

Hemodialysis is the most common method used to remove waste and hazardous products of metabolism in patients suffering from renal failure. Hundreds of thousands of people with end-stage renal disease undergo hemodialysis treatment in the United States each year. Strikingly, the 5-year survival rate for all dialysis patients is only 35%. Most of the patients succumb to cardiovascular disease that is exacerbated by the chronic induction of inflammation caused by contact of the blood with the dialysis membrane. The complement system, a strong mediator of pro-inflammatory networks, is a key contributor to such biomaterial-induced inflammation. Though only evaluated in experimental ex vivo settings, specific targeting of complement activation during hemodialysis has uncovered valuable information that points towards the therapeutic use of complement inhibitors as means to control the unwelcomed inflammatory responses and consequent pathologies in hemodialysis patients. PMID:22964235

CFHR1-Modified Neural Stem Cells Ameliorated Brain Injury in a Mouse Model of Neuromyelitis Optica Spectrum Disorders.

PubMed

Shi, Kaibin; Wang, Zhen; Liu, Yuanchu; Gong, Ye; Fu, Ying; Li, Shaowu; Wood, Kristofer; Hao, Junwei; Zhang, Guang-Xian; Shi, Fu-Dong; Yan, Yaping

2016-11-01

A major hurdle for effective stem cell therapy is ongoing inflammation in the target organ. Reconditioning the lesion microenvironment may be an effective way to promote stem cell therapy. In this study, we showed that engineered neural stem cells (NSCs) with complement factor H-related protein 1, a complement inhibitor protein, can attenuate inflammatory infiltration and immune-mediated damage of astrocytes, an important pathogenic progress in patients with neuromyelitis optica spectrum disorders. Furthermore, we demonstrated that transplantation of the complement factor H-related protein 1-modified NSCs effectively blocked the complement activation cascade and inhibited formation of the membrane attack complex, thus contributing to the protection of endogenous and transplanted NSC-differentiated astrocytes. Therefore, manipulation of the lesion microenvironment contributes to a more effective cell replacement therapeutic strategy for autoimmune diseases of the CNS. Copyright © 2016 by The American Association of Immunologists, Inc.

Urinary bactericidal activity of single doses (250, 500, 750 and 1000 mg) of levofloxacin against fluoroquinolone-resistant strains of Escherichia coli.

PubMed

Stein, Gary E; Schooley, Sharon L; Nicolau, David P

2008-10-01

Increasing resistance to fluoroquinolones in uropathogens has become a clinical concern. The purpose of this study was to analyse the urinary bactericidal activity (UBA) of levofloxacin against fluoroquinolone-resistant strains of Escherichia coli. Ten healthy adult subjects (aged 23-60 years) received single doses of levofloxacin (250, 500, 750 and 1000 mg) and then blood and urine samples were collected in intervals (0-1.5, 1.5-4, 4-8, 8-12 and 12-24h) over 24h. Both serum and urine concentrations were measured by a validated high-performance liquid chromatography assay. Bactericidal titres in urine were determined against E. coli isolates with minimum inhibitory concentrations of 0.125, 4, 8, 16, 32 and 64microg/mL for levofloxacin. The mean serum pharmacokinetic parameters for these doses of levofloxacin were similar to previously published values. The mean peak urinary concentrations (0-1.5h) were 210, 347, 620 and 536microg/mL for the 250, 500, 750 and 1000 mg dose, respectively. Each dose of levofloxacin exhibited early (0-1.5h time period) bactericidal activity in urine in virtually all subjects against E. coli strains with MICs

Molecular Imaging of Phosphorylation Events for Drug Development

PubMed Central

Chan, C. T.; Paulmurugan, R.; Reeves, R. E.; Solow-Cordero, D.; Gambhir, S. S.

2014-01-01

Purpose Protein phosphorylation mediated by protein kinases controls numerous cellular processes. A genetically encoded, generalizable split firefly luciferase (FL)-assisted complementation system was developed for noninvasive monitoring phosphorylation events and efficacies of kinase inhibitors in cell culture and in small living subjects by optical bioluminescence imaging. Procedures An Akt sensor (AST) was constructed to monitor Akt phosphorylation and the effect of different PI-3K and Akt inhibitors. Specificity of AST was determined using a non-phosphorylable mutant sensor containing an alanine substitution (ASA). Results The PI-3K inhibitor LY294002 and Akt kinase inhibitor perifosine led to temporal- and dose-dependent increases in complemented FL activities in 293T human kidney cancer cells stably expressing AST (293T/AST) but not in 293T/ASA cells. Inhibition of endogenous Akt phosphorylation and kinase activities by perifosine also correlated with increase in complemented FL activities in 293T/AST cells but not in 293T/ASA cells. Treatment of nude mice bearing 293T/AST xenografts with perifosine led to a 2-fold increase in complemented FL activities compared to that of 293T/ASA xenografts. Our system was used to screen a small chemical library for novel modulators of Akt kinase activity. Conclusion This generalizable approach for noninvasive monitoring of phosphorylation events will accelerate the discovery and validation of novel kinase inhibitors and modulators of phosphorylation events. PMID:19048345

Inhibition of lipid A-mediated type I interferon induction by bactericidal/permeability-increasing protein (BPI).

PubMed

Azuma, Masahiro; Matsuo, Aya; Fujimoto, Yukari; Fukase, Koichi; Hazeki, Kaoru; Hazeki, Osamu; Matsumoto, Misako; Seya, Tsukasa

2007-03-09

Lipopolysaccharide (LPS), a major constituent of the outer membrane of gram-negative bacteria, consists of polysaccharides and a lipid structure named lipid A. Lipid A is a typical microbial pattern molecule that serves as a ligand for Toll-like receptor 4 (TLR4). TLR4 signals the presence of lipid A to recruit adaptor molecules and induces cytokines and type I interferon (IFN) by activating transcription factor, NF-kappaB or IRF-3. Here we showed that chemically synthesized TLR4-agonistic lipid A analogues but not antagonistic lipid A activate IFN-beta promoter in TLR4-expressing HEK293 cells. The amplitude of IFN-beta promoter activation was in parallel with that of NF-kappaB. LPS-binding protein (LBP) was required for efficient IFN-beta induction in this system, and this LBP activity was antagonized by bactericidal/permeability-increasing protein (BPI). Thus, we first show that BPI blocks the TLR4 responses by exogenous administration of BPI to lipid A-sensitive cells. Although the functional mechanism whereby extra-cellular BPI modulates the intra-cellular signal pathways selected by the TLR adaptors, MyD88 and TICAM-1 (TRIF), remains unknown, we infer that the lipid A portion of LPS participates in LBP-amplified IFN-beta induction and that BPI binding to LPS leads to inhibition of the activation of NF-kappaB and IFN-beta by LPS or agonistic lipid A via TLR4 in an extrinsic mode. BPI may serve as a therapeutic potential against endotoxin shock by acting as a regulator for the MyD88- and TICAM-1 pathways in the LPS-TLR4 signaling.

Visible-Light-Induced Bactericidal Activity of a Nitrogen-Doped Titanium Photocatalyst against Human Pathogens

PubMed Central

Wong, Ming-Show; Chu, Wen-Chen; Sun, Der-Shan; Huang, Hsuan-Shun; Chen, Jiann-Hwa; Tsai, Pei-Jane; Lin, Nien-Tsung; Yu, Mei-Shiuan; Hsu, Shang-Feng; Wang, Shih-Lien; Chang, Hsin-Hou

2006-01-01

The antibacterial activity of photocatalytic titanium dioxide (TiO2) substrates is induced primarily by UV light irradiation. Recently, nitrogen- and carbon-doped TiO2 substrates were shown to exhibit photocatalytic activities under visible-light illumination. Their antibacterial activity, however, remains to be quantified. In this study, we demonstrated that nitrogen-doped TiO2 substrates have superior visible-light-induced bactericidal activity against Escherichia coli compared to pure TiO2 and carbon-doped TiO2 substrates. We also found that protein- and light-absorbing contaminants partially reduce the bactericidal activity of nitrogen-doped TiO2 substrates due to their light-shielding effects. In the pathogen-killing experiment, a significantly higher proportion of all tested pathogens, including Shigella flexneri, Listeria monocytogenes, Vibrio parahaemolyticus, Staphylococcus aureus, Streptococcus pyogenes, and Acinetobacter baumannii, were killed by visible-light-illuminated nitrogen-doped TiO2 substrates than by pure TiO2 substrates. These findings suggest that nitrogen-doped TiO2 has potential application in the development of alternative disinfectants for environmental and medical usages. PMID:16957236

Antimicrobial activity and safety evaluation of peptides isolated from the hemoglobin of chickens.

PubMed

Hu, Fengjiao; Wu, Qiaoxing; Song, Shuang; She, Ruiping; Zhao, Yue; Yang, Yifei; Zhang, Meikun; Du, Fang; Soomro, Majid Hussain; Shi, Ruihan

2016-12-05

Hemoglobin is a rich source of biological peptides. As a byproduct and even wastewater of poultry-slaughtering facilities, chicken blood is one of the most abundant source of hemoglobin. In this study, the chicken hemoglobin antimicrobial peptides (CHAP) were isolated and the antimicrobial and bactericidal activities were tested by the agarose diffusion assay, minimum inhibitory concentration (MIC) analysis, minimal bactericidal concentration (MBC) analysis, and time-dependent inhibitory and bactericidal assays. The results demonstrated that CHAP had potent and rapid antimicrobial activity against 19 bacterial strains, including 9 multidrug-resistant bacterial strains. Bacterial biofilm and NaCl permeability assays, transmission electron microscopy (TEM) and scanning electron microscopy (SEM) were further performed to detect the mechanism of its antimicrobial effect. Additionally, CHAP showed low hemolytic activity, embryo toxicity, and high stability in different temperatures and animal plasma. CHAP may have great potential for expanding production and development value in animal medication, the breeding industry and environment protection.

Determination of the bactericidal activity of different calcium hydroxide presentations on a dentin model.

PubMed

Sinan, A; Adou, A J; Rochd, T; Calas, P

2007-03-01

Calcium hydroxide placed as a temporary dressing in the root canal helps sterilize infected canals. Hycal and Roeko calcium hydroxide points are two delayed-action medications containing Ca(OH)2 recommended for root canal treatment. The purpose of this study was to test their bactericidal activity in comparison with that of C-PMCP on Streptococcus sanguins strain NCTC 7823. Artificially infected 4-mm high blocks of dentin obtained from bovine incisors were used as an experimental model (n = 192). After three days of treatment with the two antiseptics, intracanal dentin powder was collected by serial drillings and used to inoculate a culture broth. The turbidity of this broth after 24 hours showed if the bacteria were eliminated or not and was used as criterion of antiseptic efficacy. No bacterial growth was observed in the samples treated with C-PMCP. Hycal had a considerable bactericidal activity with 94% of negative cultures whereas first-generation Roeko points had no activity on the strain of S. sanguis tested.

Quantitative evaluation of dermatological antiseptics.

PubMed

Leitch, C S; Leitch, A E; Tidman, M J

2015-12-01

Topical antiseptics are frequently used in dermatological management, yet evidence for the efficacy of traditional generic formulations is often largely anecdotal. We tested the in vitro bactericidal activity of four commonly used topical antiseptics against Staphylococcus aureus, using a modified version of the European Standard EN 1276, a quantitative suspension test for evaluation of the bactericidal activity of chemical disinfectants and antiseptics. To meet the standard for antiseptic effectiveness of EN 1276, at least a 5 log10 reduction in bacterial count within 5 minutes of exposure is required. While 1% benzalkonium chloride and 6% hydrogen peroxide both achieved a 5 log10 reduction in S. aureus count, neither 2% aqueous eosin nor 1 : 10 000 potassium permanganate showed significant bactericidal activity compared with control at exposure periods of up to 1 h. Aqueous eosin and potassium permanganate may have desirable astringent properties, but these results suggest they lack effective antiseptic activity, at least against S. aureus. © 2015 British Association of Dermatologists.

Bactericidal activity against coagulase-negative staphylococci is impaired in infants receiving long-term parenteral nutrition.

PubMed

Okada, Y; Klein, N J; van Saene, H K; Webb, G; Holzel, H; Pierro, A

2000-02-01

To examine the role of total parenteral nutrition (TPN) in predisposing infants to infection caused by coagulase-negative staphylococci. Total parenteral nutrition is an important means of providing essential nutrients to newborn infants. However, its use has been associated with complications, particularly infection caused by coagulase-negative staphylococci. Recent data suggest that TPN may modulate immune function; however, reports directly indicating impaired immunity against coagulase-negative staphylococci during TPN are limited. Study 1 involved 31 infants younger than 4 months who had undergone surgery and were not receiving antibiotics; 20 were receiving TPN and 11 were receiving a normal enteral diet. An in vitro whole blood model was used to measure the host bactericidal activity against coagulase-negative staphylococci. Bacterial killing and phagocytosis were measured after a 45-minute challenge with viable coagulase-negative staphylococci. In study 2, whole blood killing and intracellular killing of coagulase-negative staphylococci were measured in five newborn infants (younger than 2 months) who were receiving long-term TPN (>10 days), five control infants receiving a normal enteral diet, and five healthy adults. In study 1, infants receiving a normal enteral diet showed a high capacity to ingest and kill coagulase-negative staphylococci. In contrast, the blood of infants receiving long-term TPN showed a reduction in coagulase-negative staphylococci phagocytosis and killing. There were significant negative linear correlations between the duration of TPN and killing of coagulase-negative staphylococci and phagocytosis of coagulase-negative staphylococci. In study 2, infants receiving long-term TPN had lower whole blood killing and intracellular killing than infants receiving a normal enteral diet and healthy adult volunteers. These data seem to indicate a neutrophil dysfunction mediated by TPN in infancy. Host defense mechanisms, including phagocytosis and killing of coagulase-negative staphylococci, are impaired during long-term TPN. The impaired bactericidal activity seems to be related to defective intracellular killing in neutrophils. These findings may explain the high rate of septicemia caused by coagulase-negative staphylococci in infants receiving TPN.

Molecular characterization of the rhesus rhadinovirus (RRV) ORF4 gene and the RRV complement control protein it encodes.

PubMed

Mark, Linda; Spiller, O Brad; Okroj, Marcin; Chanas, Simon; Aitken, Jim A; Wong, Scott W; Damania, Blossom; Blom, Anna M; Blackbourn, David J

2007-04-01

The diversity of viral strategies to modulate complement activation indicates that this component of the immune system has significant antiviral potential. One example is the Kaposi's sarcoma-associated herpesvirus (KSHV) complement control protein (KCP), which inhibits progression of the complement cascade. Rhesus rhadinovirus (RRV), like KSHV, is a member of the subfamily Gammaherpesvirinae and currently provides the only in vivo model of KSHV pathobiology in primates. In the present study, we characterized the KCP homologue encoded by RRV, RRV complement control protein (RCP). Two strains of RRV have been sequenced to date (H26-95 and 17577), and the RCPs they encode differ substantially in structure: RCP from strain H26-95 has four complement control protein (CCP) domains, whereas RCP from strain 17577 has eight CCP domains. Transcriptional analyses of the RCP gene (ORF4, referred to herein as RCP) in infected rhesus macaque fibroblasts mapped the ends of the transcripts of both strains. They revealed that H26-95 encodes a full-length, unspliced RCP transcript, while 17577 RCP generates a full-length unspliced mRNA and two alternatively spliced transcripts. Western blotting confirmed that infected cells express RCP, and immune electron microscopy disclosed this protein on the surface of RRV virions. Functional studies of RCP encoded by both RRV strains revealed their ability to suppress complement activation by the classical (antibody-mediated) pathway. These data provide the foundation for studies into the biological significance of gammaherpesvirus complement regulatory proteins in a tractable, non-human primate model.

Molecular Characterization of the Rhesus Rhadinovirus (RRV) ORF4 Gene and the RRV Complement Control Protein It Encodes▿

PubMed Central

Mark, Linda; Spiller, O. Brad; Okroj, Marcin; Chanas, Simon; Aitken, Jim A.; Wong, Scott W.; Damania, Blossom; Blom, Anna M.; Blackbourn, David J.

2007-01-01

The diversity of viral strategies to modulate complement activation indicates that this component of the immune system has significant antiviral potential. One example is the Kaposi's sarcoma-associated herpesvirus (KSHV) complement control protein (KCP), which inhibits progression of the complement cascade. Rhesus rhadinovirus (RRV), like KSHV, is a member of the subfamily Gammaherpesvirinae and currently provides the only in vivo model of KSHV pathobiology in primates. In the present study, we characterized the KCP homologue encoded by RRV, RRV complement control protein (RCP). Two strains of RRV have been sequenced to date (H26-95 and 17577), and the RCPs they encode differ substantially in structure: RCP from strain H26-95 has four complement control protein (CCP) domains, whereas RCP from strain 17577 has eight CCP domains. Transcriptional analyses of the RCP gene (ORF4, referred to herein as RCP) in infected rhesus macaque fibroblasts mapped the ends of the transcripts of both strains. They revealed that H26-95 encodes a full-length, unspliced RCP transcript, while 17577 RCP generates a full-length unspliced mRNA and two alternatively spliced transcripts. Western blotting confirmed that infected cells express RCP, and immune electron microscopy disclosed this protein on the surface of RRV virions. Functional studies of RCP encoded by both RRV strains revealed their ability to suppress complement activation by the classical (antibody-mediated) pathway. These data provide the foundation for studies into the biological significance of gammaherpesvirus complement regulatory proteins in a tractable, non-human primate model. PMID:17287274

Potassium iodide potentiates antimicrobial photodynamic inactivation mediated by Rose Bengal: in vitro and in vivo studies

NASA Astrophysics Data System (ADS)

Wen, Xiang; Zhang, Xiaoshen; Szewczyk, Grzegorz; ElHussien, Ahmed; Huang, Ying-Ying; Sarna, Tadeusz; Hamblin, Michael R.

2018-02-01

Rose Bengal (RB) is a halogenated xanthene dye that has been used to mediate antimicrobial photodynamic inactivation. While highly active against Gram-positive bacteria, RB is largely inactive in killing Gram-negative bacteria. We have discovered that addition of the non-toxic salt potassium iodide (100mM) potentiates green light (540nm)-mediated killing by up to six extra logs with Gramnegative bacteria Escherichia coli and Pseudomonas aeruginosa,Gram-positive methicillin resistant Staphylococcus aureus, and fungal yeast Candida albicans. The mechanism is proposed to be singlet oxygen addition to iodide anion to form peroxyiodide, which decomposes into radicals, finally forms hydrogen peroxide and molecular iodine. The effects of these different bactericidal species can be teased apart by comparing killing in three different scenarios: (1) cells+RB+KI are mixed together then illuminated with green light; (2) cells+RB are centrifuged then KI added then green light; (3) RB+KI+green light then cells added after light. We showed that KI could potentiate RBPDT in a mouse model of skin abrasions infected with bioluminescent P.aeruginosa.

Urease from Helicobacter pylori is inactivated by sulforaphane and other isothiocyanates.

PubMed

Fahey, Jed W; Stephenson, Katherine K; Wade, Kristina L; Talalay, Paul

2013-05-24

Infections by Helicobacter pylori are very common, causing gastroduodenal inflammation including peptic ulcers, and increasing the risk of gastric neoplasia. The isothiocyanate (ITC) sulforaphane [SF; 1-isothiocyanato-4-(methylsulfinyl)butane] derived from edible crucifers such as broccoli is potently bactericidal against Helicobacter, including antibiotic-resistant strains, suggesting a possible dietary therapy. Gastric H. pylori infections express high urease activity which generates ammonia, neutralizes gastric acidity, and promotes inflammation. The finding that SF inhibits (inactivates) urease (jack bean and Helicobacter) raised the issue of whether these properties might be functionally related. The rates of inactivation of urease activity depend on enzyme and SF concentrations and show first order kinetics. Treatment with SF results in time-dependent increases in the ultraviolet absorption of partially purified Helicobacter urease in the 260-320 nm region. This provides direct spectroscopic evidence for the formation of dithiocarbamates between the ITC group of SF and cysteine thiols of urease. The potencies of inactivation of Helicobacter urease by isothiocyanates structurally related to SF were surprisingly variable. Natural isothiocyanates closely related to SF, previously shown to be bactericidal (berteroin, hirsutin, phenethyl isothiocyanate, alyssin, and erucin), did not inactivate urease activity. Furthermore, SF is bactericidal against both urease positive and negative H. pylori strains. In contrast, some isothiocyanates such as benzoyl-ITC, are very potent urease inactivators, but are not bactericidal. The bactericidal effects of SF and other ITC against Helicobacter are therefore not obligatorily linked to urease inactivation, but may reduce the inflammatory component of Helicobacter infections. Copyright © 2013 Elsevier Inc. All rights reserved.

Photobactericidal activity of methylene blue derivatives against vancomycin-resistant Enterococcus spp.

PubMed

Wainwright, M; Phoenix, D A; Gaskell, M; Marshall, B

1999-12-01

The toxicities and phototoxicities of methylene blue and its two methylated derivatives were measured against one standard and three vancomycin-resistant pathogenic strains of Enterococcus spp. Each of the compounds was bactericidal and the derivatives exhibited photobactericidal activity on illumination at a 'light' dose of 6.3 J/cm(2) against one or more of the strains. Increased bactericidal and photobactericidal activity in the methylated derivatives is thought to be due to their higher hydrophobicities allowing greater interaction with the bacterial cell wall. In addition, the derivatives exhibited higher inherent photosensitizing efficacies.

Antibacterial activity of Psidium guajava leaf and bark against multidrug-resistant Vibrio cholerae: implication for cholera control.

PubMed

Rahim, Niaz; Gomes, Donald James; Watanabe, Haruo; Rahman, Sabita Rizwana; Chomvarin, Chariya; Endtz, Hubert Ph; Alam, Munirul

2010-07-01

In clinical cholera, a 3-day course of antibiotic complements extensive rehydration therapy by reducing stool volume, shortening the illness, and averting death. However, antibiotic therapy, which has lifesaving implications for cholera, is often hindered due to multidrug resistance in Vibrio cholerae, the cause of cholera. Crude aqueous mixture and water soluble methanol extract from leaf and bark of Psidium guajava, a tropical fruit guava of the family Myrtaceae, showed strong antibacterial activity against multidrug-resistant V. cholerae O1. The in vitro minimum inhibitory concentration of the crude aqueous mixture and water soluble methanol extract, which was bactericidal against 10(7) CFU/mL of V. cholerae was determined to be 1,250 microg/mL and 850 microg/mL, respectively. The antibacterial activity of P. guajava was stable at 100 degrees C for 15-20 min, suggesting nonprotein nature of the active component. The growth of V. cholerae in rice oral rehydration saline (ORS) was completely inhibited when 10 mg/mL (wt/vol) of crude aqueous mixture was premixed with the ORS in a ratio of 1:7 (vol. extract/vol. ORS). P. guajava, which is widely distributed in Bangladesh, thus offers great potential for use in indigenous, herbal medicine for controlling epidemics of cholera.

Role of Bactericidal Peptidoglycan Recognition Proteins in Regulating Gut Microbiota and Obesity

DTIC Science & Technology

2018-03-01

s) Pglyrps have an established role in innate immunity, bactericidal activity , and inflammatory diseases. Our new data suggest that in addition...genetic cause remains unknown. We report that deletion of the innate immunity antibacterial gene Nod2 abolishes this resistance, as Nod2−/− BALB/c mice...stimula- tion of Nod2 results in the activation of NF-κB and MAP kinase-signaling cascades and production of inflam- matory molecules and anti-microbial

An Unmutated IgM Response to the Vi Polysaccharide of Salmonella Typhi Contributes to Protective Immunity in a Murine Model of Typhoid.

PubMed

Pandya, Kalgi D; Palomo-Caturla, Isabel; Walker, Justin A; K Sandilya, Vijay; Zhong, Zhijiu; Alugupalli, Kishore R

2018-06-15

T cell-dependent B cell responses typically develop in germinal centers. Abs generated during such responses are isotype switched and have a high affinity to the Ag because of somatic hypermutation of Ab genes. B cell responses to purified polysaccharides are T cell independent and do not result in the formation of bona fide germinal centers, and the dominant Ab isotype produced during such responses is IgM with very few or no somatic mutations. Activation-induced cytidine deaminase (AID) is required for both somatic hypermutation and Ig isotype switching in humans and mice. To test the extent to which unmutated polysaccharide-specific IgM confers protective immunity, we immunized wildtype and AID -/- mice with either heat-killed Salmonella enterica serovar Typhi ( S. Typhi) or purified Vi polysaccharide (ViPS). We found that wildtype and AID -/- mice immunized with heat-killed S. Typhi generated similar anti-ViPS IgM responses. As expected, wildtype, but not AID -/- mice generated ViPS-specific IgG. However, the differences in the Ab-dependent killing of S. Typhi mediated by the classical pathway of complement activation were not statistically significant. In ViPS-immunized wildtype and AID -/- mice, the ViPS-specific IgM levels and S. Typhi bactericidal Ab titers at 7 but not at 28 d postimmunization were also comparable. To test the protective immunity conferred by these immunizations, mice were challenged with a chimeric S. Typhimurium strain expressing ViPS. Compared with their naive counterparts, immunized wildtype and AID -/- mice exhibited significantly reduced bacterial burden regardless of the route of infection. These data indicate that an unmutated IgM response to ViPS contributes to protective immunity to S. Typhi. Copyright © 2018 by The American Association of Immunologists, Inc.

Novel Fusion Protein Approach for Efficient High-Throughput Screening of Small Molecule–Mediating Protein-Protein Interactions in Cells and Living Animals

PubMed Central

Paulmurugan, Ramasamy; Gambhir, Sanjiv S.

2014-01-01

Networks of protein interactions execute many different intracellular pathways. Small molecules either synthesized within the cell or obtained from the external environment mediate many of these protein-protein interactions. The study of these small molecule–mediated protein-protein interactions is important in understanding abnormal signal transduction pathways in a variety of disorders, as well as in optimizing the process of drug development and validation. In this study, we evaluated the rapamycin-mediated interaction of the human proteins FK506-binding protein (FKBP12) rapamycin-binding domain (FRB) and FKBP12 by constructing a fusion of these proteins with a split-Renilla luciferase or a split enhanced green fluorescent protein (split-EGFP) such that complementation of the reporter fragments occurs in the presence of rapamycin. Different linker peptides in the fusion protein were evaluated for the efficient maintenance of complemented reporter activity. This system was studied in both cell culture and xenografts in living animals. We found that peptide linkers with two or four EAAAR repeat showed higher protein-protein interaction–mediated signal with lower background signal compared with having no linker or linkers with amino acid sequences GGGGSGGGGS, ACGSLSCGSF, and ACGSLSCGS-FACGSLSCGSF. A 9 ± 2-fold increase in signal intensity both in cell culture and in living mice was seen compared with a system that expresses both reporter fragments and the interacting proteins separately. In this fusion system, rapamycin induced heterodimerization of the FRB and FKBP12 moieties occurred rapidly even at very lower concentrations (0.00001 nmol/L) of rapamycin. For a similar fusion system employing split-EGFP, flow cytometry analysis showed significant level of rapamycin-induced complementation. PMID:16103094

Novel fusion protein approach for efficient high-throughput screening of small molecule-mediating protein-protein interactions in cells and living animals.

PubMed

Paulmurugan, Ramasamy; Gambhir, Sanjiv S

2005-08-15

Networks of protein interactions execute many different intracellular pathways. Small molecules either synthesized within the cell or obtained from the external environment mediate many of these protein-protein interactions. The study of these small molecule-mediated protein-protein interactions is important in understanding abnormal signal transduction pathways in a variety of disorders, as well as in optimizing the process of drug development and validation. In this study, we evaluated the rapamycin-mediated interaction of the human proteins FK506-binding protein (FKBP12) rapamycin-binding domain (FRB) and FKBP12 by constructing a fusion of these proteins with a split-Renilla luciferase or a split enhanced green fluorescent protein (split-EGFP) such that complementation of the reporter fragments occurs in the presence of rapamycin. Different linker peptides in the fusion protein were evaluated for the efficient maintenance of complemented reporter activity. This system was studied in both cell culture and xenografts in living animals. We found that peptide linkers with two or four EAAAR repeat showed higher protein-protein interaction-mediated signal with lower background signal compared with having no linker or linkers with amino acid sequences GGGGSGGGGS, ACGSLSCGSF, and ACGSLSCGSFACGSLSCGSF. A 9 +/- 2-fold increase in signal intensity both in cell culture and in living mice was seen compared with a system that expresses both reporter fragments and the interacting proteins separately. In this fusion system, rapamycin induced heterodimerization of the FRB and FKBP12 moieties occurred rapidly even at very lower concentrations (0.00001 nmol/L) of rapamycin. For a similar fusion system employing split-EGFP, flow cytometry analysis showed significant level of rapamycin-induced complementation.

Interaction of Leptospira Elongation Factor Tu with Plasminogen and Complement Factor H: A Metabolic Leptospiral Protein with Moonlighting Activities

PubMed Central

Abe, Cecília M.; Monaris, Denize; Morais, Zenaide M.; Souza, Gisele O.; Vasconcellos, Sílvio A.; Isaac, Lourdes; Abreu, Patrícia A. E.; Barbosa, Angela S.

2013-01-01

The elongation factor Tu (EF-Tu), an abundant bacterial protein involved in protein synthesis, has been shown to display moonlighting activities. Known to perform more than one function at different times or in different places, it is found in several subcellular locations in a single organism, and may serve as a virulence factor in a range of important human pathogens. Here we demonstrate that Leptospira EF-Tu is surface-exposed and performs additional roles as a cell-surface receptor for host plasma proteins. It binds plasminogen in a dose-dependent manner, and lysine residues are critical for this interaction. Bound plasminogen is converted to active plasmin, which, in turn, is able to cleave the natural substrates C3b and fibrinogen. Leptospira EF-Tu also acquires the complement regulator Factor H (FH). FH bound to immobilized EF-Tu displays cofactor activity, mediating C3b degradation by Factor I (FI). In this manner, EF-Tu may contribute to leptospiral tissue invasion and complement inactivation. To our knowledge, this is the first description of a leptospiral protein exhibiting moonlighting activities. PMID:24312361

Superior bactericidal activity of N-bromine compounds compared to their N-chlorine analogues can be reversed under protein load.

PubMed

Gottardi, W; Klotz, S; Nagl, M

2014-06-01

To investigate and compare the bactericidal activity (BA) of active bromine and chlorine compounds in the absence and presence of protein load. Quantitative killing tests against Escherichia coli and Staphylococcus aureus were performed both in the absence and in the presence of peptone with pairs of isosteric active chlorine and bromine compounds: hypochlorous and hypobromous acid (HOCl and HOBr), dichloro- and dibromoisocyanuric acid, chlorantine and bromantine (1,3-dibromo- and 1,3 dichloro-5,5-dimethylhydantoine), chloramine T and bromamine T (N-chloro- and N-bromo-4-methylbenzenesulphonamide sodium), and N-chloro- and N-bromotaurine sodium. To classify the bactericidal activities on a quantitative basis, an empirical coefficient named specific bactericidal activity (SBA), founded on the parameters of killing curves, was defined: SBA= mean log reductions/(mean exposure times x concentration) [mmol 1(-1) min (-1)]. In the absence of peptone, tests with washed micro-organisms revealed a throughout higher BA of bromine compounds with only slight differences between single substances. This was in contrast to chlorine compounds, whose killing times differed by a factor of more than four decimal powers. As a consequence, also the isosteric pairs showed according differences. In the presence of peptone, however, bromine compounds showed an increased loss of BA, which partly caused a reversal of efficacy within isosteric pairs. In medical practice, weakly oxidizing active chlorine compounds like chloramines have the highest potential as topical anti-infectives in the presence of proteinaceous material (mucous membranes, open wounds). Active bromine compounds, on the other hand, have their chance at insensitive body regions with low organic matter, for example skin surfaces. The expected protein load is one of the most important parameters for selection of a suited active halogen compound. © 2014 The Society for Applied Microbiology.

Synergistic bactericidal activity of chlorhexidine-loaded, silver-decorated mesoporous silica nanoparticles.

PubMed

Lu, Meng-Meng; Wang, Qiu-Jing; Chang, Zhi-Min; Wang, Zheng; Zheng, Xiao; Shao, Dan; Dong, Wen-Fei; Zhou, Yan-Min

2017-01-01

Combination of chlorhexidine (CHX) and silver ions could engender synergistic bactericidal effect and improve the bactericidal efficacy. It is highly desired to develop an efficient carrier for the antiseptics codelivery targeting infection foci with acidic microenvironment. In this work, monodisperse mesoporous silica nanoparticle (MSN) nanospheres were successfully developed as an ideal carrier for CHX and nanosilver codelivery through a facile and environmentally friendly method. The CHX-loaded, silver-decorated mesoporous silica nanoparticles (Ag-MSNs@CHX) exhibited a pH-responsive release manner of CHX and silver ions simultaneously, leading to synergistically antibacterial effect against both gram-positive Staphylococcus aureus and gram-negative Escherichia coli . Moreover, the effective antibacterial concentration of Ag-MSNs@CHX showed less cytotoxicity on normal cells. Given their synergistically bactericidal ability and good biocompatibility, these nanoantiseptics might have effective and broad clinical applications for bacterial infections.

Secretory phospholipase A2 in dromedary tears: a host defense against staphylococci and other gram-positive bacteria.

PubMed

Ben Bacha, Abir; Abid, Islem

2013-03-01

The best known physiologic function of secreted phospholipase A2 (sPLA2) group IIA (sPLA2-IIA) is defense against bacterial infection through hydrolytic degradation of bacterial membrane phospholipids. In fact, sPLA2-IIA effectively kills Gram-positive bacteria and to a lesser extent Gram-negative bacteria and is considered a major component of the eye's innate immune defense system. The antibacterial properties of sPLA2 have been demonstrated in rabbit and human tears. In this report, we have analyzed the bactericidal activity of dromedary tears and the subsequently purified sPLA2 on several Gram-positive bacteria. Our results showed that the sPLA2 displays a potent bactericidal activity against all the tested bacteria particularly against the Staphylococcus strains when tested in the ionic environment of tears. There is a synergic action of the sPLA2 with lysozyme when added to the bacteria culture prior to sPLA2. Interestingly, lysozyme purified from dromedary tears showed a significant bactericidal activity against Listeria monocytogene and Staphylococcus epidermidis, whereas the one purified from human tears displayed no activity against these two strains. We have also demonstrated that Ca(2+) is crucial for the activity of dromedary tear sPLA2 and to a less extent Mg(2+) ions. Given the presence of sPLA2 in tears and intestinal secretions, this enzyme may play a substantial role in innate mucosal and systemic bactericidal defenses against Gram-positive bacteria.

Colloidal metal oxide nanoparticle systems: the new promising way to prevent antibiotic resistance during treatment of local infectious processes

NASA Astrophysics Data System (ADS)

Karasenkov, Y.; Frolov, G.; Pogorelsky, I.; Latuta, N.; Gusev, A.; Kuznetsov, D.; Leont'ev, V.

2015-11-01

New bactericidal containing nanoparticles colloids for application in dentistry, maxillofacial surgery, urology, obstetrics, gynaecology, ENT, proctology have been developed. The various water colloidal nanodispersive systems of metals and oxides have been obtained by means of electric impulse - condensation (electroerosion) method. These systems are based pure elements and alloys of argentum (Ag), titanium dioxide (TiO2), iron oxide (Fe2O3), tantalum oxide (TaO), vanadium oxide (VO2), cobalt oxide (CoO), tantalum dioxide TaO2, zinc oxide (ZnO), copper oxide (CuO) and mixed suspensions of titanium, aluminium and molybdenum oxides. The research has been made on culture of dentobacterial plaque and mixed culture issued from gingival spaces. The composition of culture was identified with S.aureus, S.epidermidis and nonfermentable kinds of E.coli. The observation period lasted more than nineteen days. All solutions showed highly prolonged bactericidal activity in dilutions from the whole solution 1-20 mg/L. The bactericidal activity of powder specimen of silica containing Ag and Fe2O3 nanoparticles used as dental filling material and disintegrates of composite materials (produced by “StomaDent” CJSC) have been studied. Tested materials have long (up to 19 days and more) bactericidal activity.

Antibody-mediated rejection in kidney transplantation: a review of pathophysiology, diagnosis, and treatment options.

PubMed

Kim, Miae; Martin, Spencer T; Townsend, Keri R; Gabardi, Steven

2014-07-01

Antibody-mediated rejection (AMR), also known as B-cell-mediated or humoral rejection, is a significant complication after kidney transplantation that carries a poor prognosis. Although fewer than 10% of kidney transplant patients experience AMR, as many as 30% of these patients experience graft loss as a consequence. Although AMR is mediated by antibodies against an allograft and results in histologic changes in allograft vasculature that differ from cellular rejection, it has not been recognized as a separate disease process until recently. With an improved understanding about the importance of the development of antibodies against allografts as well as complement activation, significant advances have occurred in the treatment of AMR. The standard of care for AMR includes plasmapheresis and intravenous immunoglobulin that remove and neutralize antibodies, respectively. Agents targeting B cells (rituximab and alemtuzumab), plasma cells (bortezomib), and the complement system (eculizumab) have also been used successfully to treat AMR in kidney transplant recipients. However, the high cost of these medications, their use for unlabeled indications, and a lack of prospective studies evaluating their efficacy and safety limit the routine use of these agents in the treatment of AMR in kidney transplant recipients. © 2014 Pharmacotherapy Publications, Inc.

Chimeras of human complement C9 reveal the site recognized by complement regulatory protein CD59.

PubMed

Hüsler, T; Lockert, D H; Kaufman, K M; Sodetz, J M; Sims, P J

1995-02-24

CD59 antigen is a membrane glycoprotein that inhibits the activity of the C9 component of the C5b-9 membrane attack complex, thereby protecting human cells from lysis by human complement. The complement-inhibitory activity of CD59 is species-selective and is most effective toward C9 derived from human or other primate plasma. By contrast, rabbit C9, which can substitute for human C9 in the membrane attack complex, mediates unrestricted lysis of human cells. To identify the peptide segment of human C9 that is recognized by CD59, rabbit C9 cDNA clones were isolated, characterized, and used to construct hybrid cDNAs for expression of full-length human/rabbit C9 chimeras in COS-7 cells. All resulting chimeras were hemolytically active, when tested against chicken erythrocytes bearing C5b-8 complexes. Assays performed in the presence or absence of CD59 revealed that this inhibitor reduced the hemolytic activity of those chimeras containing human C9 sequence between residues 334-415, irrespective of whether the remainder of the protein contained human or rabbit sequence. By contrast, when this segment of C9 contained rabbit sequence, lytic activity was unaffected by CD59. These data establish that human C9 residues 334-415 contain the site recognized by CD59, and they suggest that sequence variability within this segment of C9 is responsible for the observed species-selective inhibitory activity of CD59.

Modulation of cytotoxic T lymphocyte, natural killer cell, antibody-dependent cellular cytotoxicity, and antibody-dependent complement-mediated cytotoxicity by Vernonia cinerea L. and vernolide-A in BALB/c mice via enhanced production of cytokines IL-2 and IFN-γ.

PubMed

Pratheeshkumar, Poyil; Kuttan, Girija

2012-02-01

Effect of Vernonia cinerea L. and vernolide-A on cell-mediated immune (CMI) response was studied in normal as well as tumor-bearing BALB/c mice. Administration of V. cinerea and vernolide-A significantly enhanced natural killer (NK) cell activity in both normal as well as tumor-bearing animals, and the activity was observed earlier than in tumor-bearing control animals. Antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent complement-mediated cytotoxicity (ACC) were also enhanced significantly in both normal as well as tumor-bearing animals after V. cinerea and vernolide-A administration compared with untreated control tumor-bearing animals. Extract and vernolide-A showed a significant increase in cytotoxic T lymphocyte (CTL) production in both the in vivo and in vitro models. The level of cytokines such as interleukin (IL)-2 and interferon (IFN)-γ were also enhanced by the treatment of V. cinerea and vernolide-A in both normal as well as tumor-bearing animals. This study demonstrated that V. cinerea extract and vernolide-A stimulate the CTL, NK cell, ADCC, and ADCC through enhanced secretion of IL-2 and IFN-γ.

Myasthenia gravis: the role of complement at the neuromuscular junction.

PubMed

Howard, James F

2018-01-01

Generalized myasthenia gravis (gMG) is a rare autoimmune disorder characterized by skeletal muscle weakness caused by disrupted neurotransmission at the neuromuscular junction (NMJ). Approximately 74-88% of patients with gMG have acetylcholine receptor (AChR) autoantibodies. Complement plays an important role in innate and antibody-mediated immunity, and activation and amplification of complement results in the formation of membrane attack complexes (MACs), lipophilic proteins that damage cell membranes. The role of complement in gMG has been demonstrated in animal models and patients. Studies in animals lacking specific complement proteins have confirmed that MAC formation is required to induce experimental autoimmune MG (EAMG) and NMJ damage. Complement inhibition in EAMG models can prevent disease induction and reverse its progression. Patients with anti-AChR + MG have autoantibodies and MACs present at NMJs. Damaged NMJs are associated with more severe disease, fewer AChRs, and MACs in synaptic debris. Current MG therapies do not target complement directly. Eculizumab is a humanized monoclonal antibody that inhibits cleavage of complement protein C5, preventing MAC formation. Eculizumab treatment improved symptoms compared with placebo in a phase II study in patients with refractory gMG. Direct complement inhibition could preserve NMJ physiology and muscle function in patients with anti-AChR + gMG. © 2017 The Authors. Annals of the New York Academy of Sciences published by Wiley Periodicals Inc. on behalf of The New York Academy of Sciences.

Unmasking of complements using proteinase-K in formalin fixed paraffin embedded renal biopsies.

PubMed

Nada, R; Kumar, A; Kumar, V G; Gupta, K L; Joshi, K

2016-01-01

Renal biopsy interpretation requires histopathology, direct immunofluorescence (DIF) and electron microscopy. Formalin-fixed, paraffin-embedded tissue (FFPE) sent for light microscopy can be used for DIF after antigen retrieval. However, complement staining has not been satisfactory. We standardized DIF using proteinase-K for antigen retrieval in FFPE renal biopsies. A pilot study was conducted on known cases of membranous glomerulonephritis (MGN), membranoproliferative type-1 (MPGN-1), immunoglobulin A nephropathy (IgAN), and anti-glomerular basement disease (anti-GBM). Immunofluorescence panel included fluorescein isothiocyanate (FITC) conjugated IgG, IgA, IgM, complements (C3 and C1q), light chains (kappa, lambda) and fibrinogen antibodies. After standardization of the technique, 75 renal biopsies and 43 autopsies cases were stained. Out of 43 autopsy cases, immune-complex mediated glomerulonephritis (GN) was confirmed in 18 cases (Lupus nephritis-11, IgAN-6, MGN-1), complement-mediated dense deposit disease (DDD-1) and monoclonal diseases in 4 cases (amyloidosis-3, cast nephropathy-1). Immune-mediated injury was excluded in 17 cases (focal segmental glomerulosclerosis -3, crescentic GN-6 [pauci-immune-3, anti-GBM-3], thrombotic microangiopathy-5, atherosclerosis-3). Renal biopsies (n-75) where inadequate or no frozen sample was available; this technique classified 52 mesangiocapillary pattern as MPGN type-1-46, DDD-2 and (C3GN-4). Others were diagnosed as IgAN-3, lupus nephritis-2, MGN-4, diffuse proliferative glomerulonephritis (DPGN)-1, Non-IC crescentic GN-1, monoclonal diseases-3. In nine cases, DIF on FFPE tissue could not help in making diagnosis. Proteinase-K enzymatic digestion of FFPE renal biopsies can unmask complements (both C3 and C1q) in immune-complexes mediated and complement-mediated diseases. This method showed good results on autopsy tissues archived for as long as 15 years.

Antagonism between Bacteriostatic and Bactericidal Antibiotics Is Prevalent

PubMed Central

Lázár, Viktória; Papp, Balázs; Arnoldini, Markus; Abel zur Wiesch, Pia; Busa-Fekete, Róbert; Fekete, Gergely; Pál, Csaba; Ackermann, Martin; Bonhoeffer, Sebastian

2014-01-01

Combination therapy is rarely used to counter the evolution of resistance in bacterial infections. Expansion of the use of combination therapy requires knowledge of how drugs interact at inhibitory concentrations. More than 50 years ago, it was noted that, if bactericidal drugs are most potent with actively dividing cells, then the inhibition of growth induced by a bacteriostatic drug should result in an overall reduction of efficacy when the drug is used in combination with a bactericidal drug. Our goal here was to investigate this hypothesis systematically. We first constructed time-kill curves using five different antibiotics at clinically relevant concentrations, and we observed antagonism between bactericidal and bacteriostatic drugs. We extended our investigation by performing a screen of pairwise combinations of 21 different antibiotics at subinhibitory concentrations, and we found that strong antagonistic interactions were enriched significantly among combinations of bacteriostatic and bactericidal drugs. Finally, since our hypothesis relies on phenotypic effects produced by different drug classes, we recreated these experiments in a microfluidic device and performed time-lapse microscopy to directly observe and quantify the growth and division of individual cells with controlled antibiotic concentrations. While our single-cell observations supported the antagonism between bacteriostatic and bactericidal drugs, they revealed an unexpected variety of cellular responses to antagonistic drug combinations, suggesting that multiple mechanisms underlie the interactions. PMID:24867991

Algerian propolis extracts: Chemical composition, bactericidal activity and in vitro effects on gilthead seabream innate immune responses.

PubMed

Soltani, El-Khamsa; Cerezuela, Rebeca; Charef, Noureddine; Mezaache-Aichour, Samia; Esteban, Maria Angeles; Zerroug, Mohamed Mihoub

2017-03-01

Propolis has been used as a medicinal agent for centuries. The chemical composition of four propolis samples collected from four locations of the Sétif region, Algeria, using gas chromatography-mass spectrometry was determined. More than 20 compounds and from 30 to 35 compounds were identified in the aqueous and ethanolic extracts, respectively. Furthermore, the antimicrobial activity of the propolis extracts against two marine pathogenic bacteria was evaluated. Finally, the in vitro effects of propolis on gilthead seabream (Sparus aurata L.) leucocyte activities were measured. The bactericidal activity of ethanolic extracts was very high against Shewanella putrefaciens, average against Photobacterium damselae and very low against Vibrio harveyi. The lowest bactericidal activity was always that found for the aqueous extracts. When the viability of gilthead seabream head-kidney leucocytes was measured after 30 min' incubation with the different extracts, both the ethanolic and aqueous extracts of one of the propolis samples (from Babor) and the aqueous extract of another (from Ain-Abbassa) provoked a significant decrease in cell viability when used at concentrations of 100 and 200 μg ml -1 . Furthermore, significant inhibitory effects were recorded on leucocyte respiratory burst activity when isolated leucocytes where preincubated with the extracts. This effect was dose-dependent in all cases except when extracts from a third propolis sample (from Boutaleb) were used. Our findings suggest that some of Algerian propolis extracts have bactericidal activity against important bacterial pathogens in seabream and significantly modulate in vitro leucocyte activities, confirming their potential as a source of new natural biocides and/or immunomodulators in aquaculture practice. Copyright © 2017 Elsevier Ltd. All rights reserved.

Synthesis, anti-inflammatory, bactericidal activities and docking studies of novel 1,2,3-triazoles derived from ibuprofen using click chemistry.

PubMed

Angajala, Kishore Kumar; Vianala, Sunitha; Macha, Ramesh; Raghavender, M; Thupurani, Murali Krishna; Pathi, P J

2016-01-01

Nonsteroidal anti-inflammatory drugs are of vast therapeutic benefit in the treatment of different types of inflammatory conditions. 1,2,3-Triazoles and their derivatives have a wide range of applications as anti-bacterial, anti-fungal, anti-tubercular, cytostatic, anti-HIV, anti-allergic, anti-neoplastic and anti-inflammatory (AI) agents. Considering the individual biological and medicinal importance of ibuprofen and 1,2,3-triazoles, we wanted to explore novel chemical entities based on ibuprofen and triazole moieties towards their biological significance. Click chemistry has utilized as an ideal strategy to prepare novel ibuprofen-based 1,4-disubstituted 1,2,3-triazole containing molecules. These compounds were screened for their in vivo AI activity, among all the synthesized analogues 13o was shown potent effect than the reference AI drug ibuprofen at the same concentration (10 mg/kg body weight). Compounds 13l, 13g, 13c, 13k, 13i, 13n, 13m and 13j were shown significant AI activity. These triazole analogues were also screened for their bactericidal profile. Compounds 13c, 13i, 13l and 13o exhibited considerable bactericidal activity against gram positive and gram negative strains. In addition to this, molecular docking studies were also carried out into cyclooxygenase-2 active site to predict the affinity and orientation of these novel compounds (13a-q). In summary, we have designed and synthesized 1,2,3-triazole analogues of ibuprofen in good yields using Click chemistry approach. AI and bactericidal activities of these compounds were evaluated and shown remarkable results.

Complement Interaction with Trypanosomatid Promastigotes in Normal Human Serum

PubMed Central

Domínguez, Mercedes; Moreno, Inmaculada; López-Trascasa, Margarita; Toraño, Alfredo

2002-01-01

In normal human serum (NHS), axenic promastigotes of Crithidia, Phytomonas, and Leishmania trigger complement activation, and from 1.2 to 1.8 × 105 C3 molecules are deposited per promastigote within 2.5 min. In Leishmania, promastigote C3 binding capacity remains constant during in vitro metacyclogenesis. C3 deposition on promastigotes activated through the classical complement pathway reaches a 50% maximum after ∼50 s, and represents >85% of total C3 bound. In C1q- and C2-deficient human sera, promastigotes cannot activate the classical pathway (CP) unless purified C1q or C2 factors, respectively, are supplemented, demonstrating a requirement for CP factor in promastigote C3 opsonization. NHS depleted of natural anti-Leishmania antibodies cannot trigger promastigote CP activation, but IgM addition restores C3 binding. Furthermore, Leishmania binds natural antibodies in ethylenediaminetetracetic acid (EDTA)-treated NHS; after EDTA removal, promastigote-bound IgM triggers C3 deposition in natural antibody-depleted NHS. Serum collectins and pentraxins thus do not participate significantly in NHS promastigote C3 opsonization. Real-time kinetic analysis of promastigote CP-mediated lysis indicates that between 85–95% of parasites are killed within 2.5 min of serum contact. These data indicate that successful Leishmania infection in man must immediately follow promastigote transmission, and that Leishmania evasion strategies are shaped by the selective pressure exerted by complement. PMID:11854358

Activation of airway epithelial bitter taste receptors by Pseudomonas aeruginosa quinolones modulates calcium, cyclic-AMP, and nitric oxide signaling.

PubMed

Freund, Jenna R; Mansfield, Corrine J; Doghramji, Laurel J; Adappa, Nithin D; Palmer, James N; Kennedy, David W; Reed, Danielle R; Jiang, Peihua; Lee, Robert J

2018-05-10

Bitter taste receptors (T2Rs), discovered in many tissues outside the tongue, have recently become potential therapeutic targets. We showed previously that airway epithelial cells express several T2Rs that activate innate immune responses that may be important for treatment of airway diseases such as chronic rhinosinusitis. It is imperative to more clearly understand what compounds activate airway T2Rs as well as their full range of functions. T2R isoforms in airway motile cilia (T2Rs 4, 14, 16, and 38) produce bactericidal levels of nitric oxide (NO) that also increase ciliary beating, promoting clearance of mucus and trapped pathogens. Bacterial quorum-sensing acyl-homoserine lactones (AHLs) activate T2Rs and stimulate these responses in primary airway cells.  Quinolones are another type of quorum sensing molecule used by Pseudomonas aeruginosa.  To elucidate if bacterial quinolones activate airway T2Rs, we analyzed calcium, cAMP, and NO dynamics using a combination of fluorescent indicator dyes and FRET-based protein biosensors.  T2R-transfected HEK293T cells, several lung epithelial cell lines, and primary sinonasal cells grown and differentiated at air-liquid interface were tested with 2-heptyl-3-hydroxy-4-quinolone (known as Pseudomonas quinolone signal; PQS), 2,4-dihydroxyquinolone (DHQ), and 4-hydroxy-2-heptylquinolone (HHQ). In HEK293T cells, PQS activated T2R4, 16, and 38 while HHQ activated T2R14.  DHQ had no effect.  PQS and HHQ increased calcium and decreased both baseline and stimulated cAMP levels in cultured and primary airway cells.  In primary cells, PQS and HHQ activated levels of NO synthesis previously shown to be bactericidal. This study suggests airway T2R-mediated immune responses are activated by bacterial quinolones as well as AHLs. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.

Synthesis and Complete Antimicrobial Characterization of CEOBACTER, an Ag-Based Nanocomposite

PubMed Central

Vasquez-Peña, M.; Raymond-Herrera, O.; Villavicencio-García, H.; Petranovskii, V.; Vazquez-Duhalt, R.; Huerta-Saquero, A.

2016-01-01

The antimicrobial activity of silver nanoparticles (AgNPs) is currently used as an alternative disinfectant with diverse applications, ranging from decontamination of aquatic environments to disinfection of medical devices and instrumentation. However, incorporation of AgNPs to the environment causes collateral damage that should be avoided. In this work, a novel Ag-based nanocomposite (CEOBACTER) was successfully synthetized. It showed excellent antimicrobial properties without the spread of AgNPs into the environment. The complete CEOBACTER antimicrobial characterization protocol is presented herein. It is straightforward and reproducible and could be considered for the systematic characterization of antimicrobial nanomaterials. CEOBACTER showed minimal bactericidal concentration of 3 μg/ml, bactericidal action time of 2 hours and re-use capacity of at least five times against E. coli cultures. The bactericidal mechanism is the release of Ag ions. CEOBACTER displays potent bactericidal properties, long lifetime, high stability and re-use capacity, and it does not dissolve in the solution. These characteristics point to its potential use as a bactericidal agent for decontamination of aqueous environments. PMID:27824932

Is Research Mediating the Relationship between Teaching Experience and Student Satisfaction?

ERIC Educational Resources Information Center

Berbegal-Mirabent, Jasmina; Mas-Machuca, Marta; Marimon, Frederic

2018-01-01

Universities must ensure that academic staff are qualified and competent for performing their job. Teaching and research are two key activities in which lecturers should excel. While some authors suggest teaching and research complement each other and positively influence student satisfaction, some others point to a rivalry effect between the two.…

A Regulatory Role for Src Homology 2 Domain–Containing Inositol 5′-Phosphatase (Ship) in Phagocytosis Mediated by Fcγ Receptors and Complement Receptor 3 (αMβ2; Cd11b/Cd18)

PubMed Central

Cox, Dianne; Dale, Benjamin M.; Kashiwada, Masaki; Helgason, Cheryl D.; Greenberg, Steven

2001-01-01

The Src homology 2 domain–containing inositol 5′-phosphatase (SHIP) is recruited to immunoreceptor tyrosine-based inhibition motif (ITIM)–containing proteins, thereby suppressing phosphatidylinositol 3-kinase (PI 3-kinase)–dependent pathways. The role of SHIP in phagocytosis, a PI 3-kinase–dependent pathway, is unknown. Overexpression of SHIP in macrophages led to an inhibition of phagocytosis mediated by receptors for the Fc portion of IgG (FcγRs). In contrast, macrophages expressing catalytically inactive SHIP or lacking SHIP expression demonstrated enhanced phagocytosis. To determine whether SHIP regulates phagocytosis mediated by receptors that are not known to recruit ITIMs, we determined the effect of SHIP expression on complement receptor 3 (CR3; CD11b/CD18; αMβ2)–dependent phagocytosis. Macrophages overexpressing SHIP demonstrated impaired CR3-mediated phagocytosis, whereas macrophages expressing catalytically inactive SHIP demonstrated enhanced phagocytosis. CR3-mediated phagocytosis in macrophages derived from SHIP−/− mice was up to 2.5 times as efficient as that observed in macrophages derived from littermate controls. SHIP was localized to FcγR- and CR3-containing phagocytic cups and was recruited to the cytoskeleton upon clustering of CR3. In a transfected COS cell model of activation-independent CR3-mediated phagocytosis, catalytically active but not inactive SHIP also inhibited phagocytosis. We conclude that PI 3-kinase(s) and SHIP regulate multiple forms of phagocytosis and that endogenous SHIP plays a role in modulating β2 integrin outside-in signaling. PMID:11136821

Correlations between phytohemagglutinin response and leukocyte profile, and bactericidal capacity in a wild rodent.

PubMed

Zhang, Zhiqiang; Zhao, Zhijun

2015-05-01

Phytohemagglutinin (PHA)-induced swelling is widely used to investigate cell-mediated and innate immunity across different vertebrate taxa. However, its physiological mechanism is still an open question due to the complexity of the involved immune components. In the present study, we measured the synchronous variations of PHA response, the proportion of different subtypes of leukocytes, as well as serum bactericidal capacity in circulation blood at 6, 12 and 24 h after PHA versus PBS injection in striped hamster, Cricetulus barabensis. First, the results showed that PHA responses reached a peak at 6 h postinjection, then sharply declined at 12 h and 24 h postinjection. Serum bactericidal capacity was higher at 6 h and 12 h than at 24 h. The proportion of different subtypes of leukocytes, as well as the ratio of neutrophils to lymphocytes did not display significant changes across different time points. Second, PHA response was positively correlated with the proportion of neutrophils and serum bactericidal capacity. The proportion of monocytes was negatively correlated with that of eosinophils and neutrophils. The proportion of basophils was negatively correlated with that of lymphocytes. Our results indicate that earlier enhanced PHA response is important for the striped hamster to cope with changing environmental conditions due to its small body mass, and the increased components of innate immunity in circulation blood may contribute to the enhancement of PHA swelling response. © 2015 International Society of Zoological Sciences, Institute of Zoology/Chinese Academy of Sciences and Wiley Publishing Asia Pty Ltd.

Novel Scabies Mite Serpins Inhibit the Three Pathways of the Human Complement System

PubMed Central

Mika, Angela; Reynolds, Simone L.; Mohlin, Frida C.; Willis, Charlene; Swe, Pearl M.; Pickering, Darren A.; Halilovic, Vanja; Wijeyewickrema, Lakshmi C.; Pike, Robert N.; Blom, Anna M.; Kemp, David J.; Fischer, Katja

2012-01-01

Scabies is a parasitic infestation of the skin by the mite Sarcoptes scabiei that causes significant morbidity worldwide, in particular within socially disadvantaged populations. In order to identify mechanisms that enable the scabies mite to evade human immune defenses, we have studied molecules associated with proteolytic systems in the mite, including two novel scabies mite serine protease inhibitors (SMSs) of the serpin superfamily. Immunohistochemical studies revealed that within mite-infected human skin SMSB4 (54 kDa) and SMSB3 (47 kDa) were both localized in the mite gut and feces. Recombinant purified SMSB3 and SMSB4 did not inhibit mite serine and cysteine proteases, but did inhibit mammalian serine proteases, such as chymotrypsin, albeit inefficiently. Detailed functional analysis revealed that both serpins interfered with all three pathways of the human complement system at different stages of their activation. SMSB4 inhibited mostly the initial and progressing steps of the cascades, while SMSB3 showed the strongest effects at the C9 level in the terminal pathway. Additive effects of both serpins were shown at the C9 level in the lectin pathway. Both SMSs were able to interfere with complement factors without protease function. A range of binding assays showed direct binding between SMSB4 and seven complement proteins (C1, properdin, MBL, C4, C3, C6 and C8), while significant binding of SMSB3 occurred exclusively to complement factors without protease function (C4, C3, C8). Direct binding was observed between SMSB4 and the complement proteases C1s and C1r. However no complex formation was observed between either mite serpin and the complement serine proteases C1r, C1s, MASP-1, MASP-2 and MASP-3. No catalytic inhibition by either serpin was observed for any of these enzymes. In summary, the SMSs were acting at several levels mediating overall inhibition of the complement system and thus we propose that they may protect scabies mites from complement-mediated gut damage. PMID:22792350

The complement system and toll-like receptors as integrated players in the pathophysiology of atherosclerosis.

PubMed

Hovland, Anders; Jonasson, Lena; Garred, Peter; Yndestad, Arne; Aukrust, Pål; Lappegård, Knut T; Espevik, Terje; Mollnes, Tom E

2015-08-01

Despite recent medical advances, atherosclerosis is a global burden accounting for numerous deaths and hospital admissions. Immune-mediated inflammation is a major component of the atherosclerotic process, but earlier research focus on adaptive immunity has gradually switched towards the role of innate immunity. The complement system and toll-like receptors (TLRs), and the crosstalk between them, may be of particular interest both with respect to pathogenesis and as therapeutic targets in atherosclerosis. Animal studies indicate that inhibition of C3a and C5a reduces atherosclerosis. In humans modified LDL-cholesterol activate complement and TLRs leading to downstream inflammation, and histopathological studies indicate that the innate immune system is present in atherosclerotic lesions. Moreover, clinical studies have demonstrated that both complement and TLRs are upregulated in atherosclerotic diseases, although interventional trials have this far been disappointing. However, based on recent research showing an intimate interplay between complement and TLRs we propose a model in which combined inhibition of both complement and TLRs may represent a potent anti-inflammatory therapeutic approach to reduce atherosclerosis. Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Expanding the potential of NAI-107 for treating serious ESKAPE pathogens: synergistic combinations against Gram-negatives and bactericidal activity against non-dividing cells

PubMed Central

Brunati, Cristina; Thomsen, Thomas T; Gaspari, Eleonora; Maffioli, Sonia; Sosio, Margherita; Jabes, Daniela; Løbner-Olesen, Anders; Donadio, Stefano

2018-01-01

Abstract Objectives To characterize NAI-107 and related lantibiotics for their in vitro activity against Gram-negative pathogens, alone or in combination with polymyxin, and against non-dividing cells or biofilms of Staphylococcus aureus. NAI-107 was also evaluated for its propensity to select or induce self-resistance in Gram-positive bacteria. Methods We used MIC determinations and chequerboard experiments to establish the antibacterial activity of the examined compounds against target microorganisms. Time–kill assays were used to evaluate killing of exponential and stationary-phase cells. The effects on biofilms (growth inhibition and biofilm eradication) were evaluated using biofilm-coated pegs. The frequency of spontaneous resistant mutants was evaluated by either direct plating or by continuous sub-culturing at 0.5 × MIC levels, followed by population analysis profiles. Results The results showed that NAI-107 and its brominated variant are highly active against Neisseria gonorrhoeae and some other fastidious Gram-negative pathogens. Furthermore, all compounds strongly synergized with polymyxin against Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa, and showed bactericidal activity. Surprisingly, NAI-107 alone was bactericidal against non-dividing A. baumannii cells. Against S. aureus, NAI-107 and related lantibiotics showed strong bactericidal activity against dividing and non-dividing cells. Activity was also observed against S. aureus biofilms. As expected for a lipid II binder, no significant resistance to NAI-107 was observed by direct plating or serial passages. Conclusions Overall, the results of the current work, along with previously published results on the efficacy of NAI-107 in experimental models of infection, indicate that this lantibiotic represents a promising option in addressing the serious threat of antibiotic resistance. PMID:29092042

Expanding the potential of NAI-107 for treating serious ESKAPE pathogens: synergistic combinations against Gram-negatives and bactericidal activity against non-dividing cells.

PubMed

Brunati, Cristina; Thomsen, Thomas T; Gaspari, Eleonora; Maffioli, Sonia; Sosio, Margherita; Jabes, Daniela; Løbner-Olesen, Anders; Donadio, Stefano

2018-02-01

To characterize NAI-107 and related lantibiotics for their in vitro activity against Gram-negative pathogens, alone or in combination with polymyxin, and against non-dividing cells or biofilms of Staphylococcus aureus. NAI-107 was also evaluated for its propensity to select or induce self-resistance in Gram-positive bacteria. We used MIC determinations and chequerboard experiments to establish the antibacterial activity of the examined compounds against target microorganisms. Time-kill assays were used to evaluate killing of exponential and stationary-phase cells. The effects on biofilms (growth inhibition and biofilm eradication) were evaluated using biofilm-coated pegs. The frequency of spontaneous resistant mutants was evaluated by either direct plating or by continuous sub-culturing at 0.5 × MIC levels, followed by population analysis profiles. The results showed that NAI-107 and its brominated variant are highly active against Neisseria gonorrhoeae and some other fastidious Gram-negative pathogens. Furthermore, all compounds strongly synergized with polymyxin against Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa, and showed bactericidal activity. Surprisingly, NAI-107 alone was bactericidal against non-dividing A. baumannii cells. Against S. aureus, NAI-107 and related lantibiotics showed strong bactericidal activity against dividing and non-dividing cells. Activity was also observed against S. aureus biofilms. As expected for a lipid II binder, no significant resistance to NAI-107 was observed by direct plating or serial passages. Overall, the results of the current work, along with previously published results on the efficacy of NAI-107 in experimental models of infection, indicate that this lantibiotic represents a promising option in addressing the serious threat of antibiotic resistance. © The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.

Antimicrobial activity of polyhexamethylene guanidine phosphate in comparison to chlorhexidine using the quantitative suspension method.

PubMed

Vitt, A; Sofrata, A; Slizen, V; Sugars, R V; Gustafsson, A; Gudkova, E I; Kazeko, L A; Ramberg, P; Buhlin, K

2015-07-17

Polyhexamethylene guanidine phosphate (PHMG-P) belongs to the polymeric guanidine family of biocides and contains a phosphate group, which may confer better solubility, a detoxifying effect and may change the kinetics and dynamics of PHMG-P interactions with microorganisms. Limited data regarding PHMG-P activity against periodontopathogenic and cariogenic microorganisms necessitates studies in this area. Aim is to evaluate polyhexamethylene guanidine phosphate antimicrobial activity in comparison to chlorhexidine. Quantitative suspension method was used enrolling Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli and Candida albicans, Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Streptococcus mutans and Lactobacillus acidophilus. Both tested antiseptics at their clinically-used concentrations, of 0.2% (w/v) and 1% (w/v), correspondingly provided swift bactericidal effects against S. aureus, P. aeruginosa, E. coli and C. albicans, A. actinomycetemcomitans and P. gingivalis with reduction factors higher than 6.0. Diluted polyhexamethylene guanidine phosphate and chlorhexidine to 0.05% continued to display anti-bacterial activity and decreased titers of standard quality control, periopathogens to below 1.0 × 10(3) colony forming units/ml, albeit requiring prolonged exposure time. To achieve a bactericidal effect against S. mutans, both antiseptics at all concentrations required a longer exposure time. We found that a clinically-used 1% of polyhexamethylene guanidine phosphate concentration did not have activity against L. acidophilus. High RF of polyhexamethylene guanidine phosphate and retention of bactericidal effects, even at 0.05%, support the use of polyhexamethylene guanidine phosphate as a biocide with sufficient anti-microbial activity against periopathogens. Polyhexamethylene guanidine phosphate displayed bactericidal activity against periopathogens and S. mutans and could potentially be applied in the management of oral diseases.

Early Components of the Complement Classical Activation Pathway in Human Systemic Autoimmune Diseases

PubMed Central

Lintner, Katherine E.; Wu, Yee Ling; Yang, Yan; Spencer, Charles H.; Hauptmann, Georges; Hebert, Lee A.; Atkinson, John P.; Yu, C. Yung

2016-01-01

The complement system consists of effector proteins, regulators, and receptors that participate in host defense against pathogens. Activation of the complement system, via the classical pathway (CP), has long been recognized in immune complex-mediated tissue injury, most notably systemic lupus erythematosus (SLE). Paradoxically, a complete deficiency of an early component of the CP, as evidenced by homozygous genetic deficiencies reported in human, are strongly associated with the risk of developing SLE or a lupus-like disease. Similarly, isotype deficiency attributable to a gene copy-number (GCN) variation and/or the presence of autoantibodies directed against a CP component or a regulatory protein that result in an acquired deficiency are relatively common in SLE patients. Applying accurate assay methodologies with rigorous data validations, low GCNs of total C4, and heterozygous and homozygous deficiencies of C4A have been shown as medium to large effect size risk factors, while high copy numbers of total C4 or C4A as prevalent protective factors, of European and East-Asian SLE. Here, we summarize the current knowledge related to genetic deficiency and insufficiency, and acquired protein deficiencies for C1q, C1r, C1s, C4A/C4B, and C2 in disease pathogenesis and prognosis of SLE, and, briefly, for other systemic autoimmune diseases. As the complement system is increasingly found to be associated with autoimmune diseases and immune-mediated diseases, it has become an attractive therapeutic target. We highlight the recent developments and offer a balanced perspective concerning future investigations and therapeutic applications with a focus on early components of the CP in human systemic autoimmune diseases. PMID:26913032

[Antibacterial activity of sulopenem, a new parenteral penem antibiotic].

PubMed

Inoue, E; Komoto, E; Taniyama, Y; Mitsuhashi, S

1996-04-01

Sulopenem, a new penem antibiotic, was compared with other antibiotics with regard to in vitro antibacterial and bactericidal activities, stabilization against beta-lactamases, and effect on the release of lipopolysaccharide from Gram-negative bacteria. The results are summarized as follows. 1. Sulopenem showed more potent activities than other antibiotics against both Gram-positive and Gram-negative bacteria except Pseudomonas aeruginosa. 2. Sulopenem showed potent bactericidal activities (MIC/MBC) against both Gram-positive and Gram-negative bacteria. Time kill studies against Staphylococcus aureus, Escherichia coli, Enterobacter cloacae and Citrobacter freundii showed potent bactericidal activities of sulopenem. 3. Sulopenem was found to possess a stronger activity than other antibiotics against beta-lactamase-producing strains except P. aeruginosa and Stenotrophomonas maltophilia. 4. In particular, sulopenem was found to be more stable to the hydrolysis by various beta-lactamases produced by Gram-negative bacteria than any other antibiotics tested. Vmax/Km values of sulopenem were smaller than those of cefotiam for all tested beta-lactamases, which reflected a broad antibacterial spectrum of sulopenem. 5. E. coli ML4707 exposed to sulopenem and imipenem released less endotoxin than did controls at all concentration ranges tested. In contrast, the strain exposed to ceftazidime at bacteriostatic concentrations released a large amount of endotoxin.

Phytosynthesis of Silver Nanoparticles Using Myrtus communis L. Leaf Extract and Investigation of Bactericidal Activity

NASA Astrophysics Data System (ADS)

Ajdari, M. R.; Tondro, G. H.; Sattarahmady, N.; Parsa, A.; Heli, H.

2017-12-01

Silver nanoparticles have been synthesized using only Myrtus communis L. leaf extract by a facile procedure without other reagents. The extract played the roles of both reducing and capping agent. The nanoparticles were characterized using field-emission scanning microscopy, and remained stable for at least 3 weeks. Antibacterial activity of the nanoparticles was evaluated toward Escherichia coli, Bacillus subtilis, Pseudomonas aeruginosa, Staphylococcus aureus, methicillin-resistant Staphylococcus aureus, and Enterococcus faecalis based on inhibition zone disk diffusion assays. The minimum inhibitory and bactericidal concentrations of the nanoparticles were obtained. Mechanisms for the antibacterial activity were proposed.

Effect of primycin on growth-arrested cultures and cell integrity of Staphylococcus aureus.

PubMed

Feiszt, Péter; Schneider, György; Emődy, Levente

2017-06-01

Bactericidal effect against non-dividing bacteria is a very advantageous, but rare characteristic among antimicrobial agents, mostly possessed by those affecting the cell membrane. These kinds of agents can kill bacterial cells without lysis. We assessed these characteristics on primycin, a topical anti-staphylococcal agent highly effective against prevalent multiresistant strains, as it also acts on the cell membrane. In time-kill studies, primycin preserved its bactericidal activity against growth-arrested Staphylococcus aureus cultures. The bactericidal action was slower against growth-arrested cultures compared to the exponentially growing ones to different extents depending on the manner of arrest. The bactericidal effect was less influenced by stringent response and by protein synthesis inhibition, proving that it does not depend on metabolic activity. In contrast, uncoupling of the membrane potential predominantly slowed, and low temperature almost stopped killing of bacteria. In consideration of published data, these facts suggest that the antibacterial action of primycin involves disrupting of the membrane potential, and is predominantly influenced by the membrane fluidity. Optical density measurements and transmission electron microscopy verified that primycin kills bacterial cells without lysis. These results reveal favorable characteristics of primycin and point to, and broaden the knowledge on its membrane-targeted effect.

Complement activation on B lymphocytes opsonized with rituximab or ofatumumab produces substantial changes in membrane structure preceding cell lysis.

PubMed

Beum, Paul V; Lindorfer, Margaret A; Beurskens, Frank; Stukenberg, P Todd; Lokhorst, Henk M; Pawluczkowycz, Andrew W; Parren, Paul W H I; van de Winkel, Jan G J; Taylor, Ronald P

2008-07-01

Binding of the CD20 mAb rituximab (RTX) to B lymphocytes in normal human serum (NHS) activates complement (C) and promotes C3b deposition on or in close proximity to cell-bound RTX. Based on spinning disk confocal microscopy analyses, we report the first real-time visualization of C3b deposition and C-mediated killing of RTX-opsonized B cells. C activation by RTX-opsonized Daudi B cells induces rapid membrane blebbing and generation of long, thin structures protruding from cell surfaces, which we call streamers. Ofatumumab, a unique mAb that targets a distinct binding site (the small loop epitope) of the CD20 Ag, induces more rapid killing and streaming on Daudi cells than RTX. In contrast to RTX, ofatumumab promotes streamer formation and killing of ARH77 cells and primary B cells from patients with chronic lymphocytic leukemia. Generation of streamers requires C activation; no streaming occurs in media, NHS-EDTA, or in sera depleted of C5 or C9. Streamers can be visualized in bright field by phase imaging, and fluorescence-staining patterns indicate they contain membrane lipids and polymerized actin. Streaming also occurs if cells are reacted in medium with bee venom melittin, which penetrates cells and forms membrane pores in a manner similar to the membrane-attack complex of C. Structures similar to streamers are demonstrable when Ab-opsonized sheep erythrocytes (non-nucleated cells) are reacted with NHS. Taken together, our findings indicate that the membrane-attack complex is a key mediator of streaming. Streamer formation may, thus, represent a membrane structural change that can occur shortly before complement-induced cell death.

Impact of Bacterial Membrane Fatty Acid Composition on the Failure of Daptomycin To Kill Staphylococcus aureus.

PubMed

Boudjemaa, Rym; Cabriel, Clément; Dubois-Brissonnet, Florence; Bourg, Nicolas; Dupuis, Guillaume; Gruss, Alexandra; Lévêque-Fort, Sandrine; Briandet, Romain; Fontaine-Aupart, Marie-Pierre; Steenkeste, Karine

2018-07-01

Daptomycin is a last-resort membrane-targeting lipopeptide approved for the treatment of drug-resistant staphylococcal infections, such as bacteremia and implant-related infections. Although cases of resistance to this antibiotic are rare, increasing numbers of clinical, in vitro , and animal studies report treatment failure, notably against Staphylococcus aureus The aim of this study was to identify the features of daptomycin and its target bacteria that lead to daptomycin treatment failure. We show that daptomycin bactericidal activity against S. aureus varies significantly with the growth state and strain, according to the membrane fatty acid composition. Daptomycin efficacy as an antibiotic relies on its ability to oligomerize within membranes and form pores that subsequently lead to cell death. Our findings ascertain that daptomycin interacts with tolerant bacteria and reaches its membrane target, regardless of its bactericidal activity. However, the final step of pore formation does not occur in cells that are daptomycin tolerant, strongly suggesting that it is incapable of oligomerization. Importantly, membrane fatty acid contents correlated with poor daptomycin bactericidal activity, which could be manipulated by fatty acid addition. In conclusion, daptomycin failure to treat S. aureus is not due to a lack of antibiotic-target interaction, but is driven by its capacity to form pores, which depends on membrane composition. Manipulation of membrane fluidity to restore S. aureus daptomycin bactericidal activity in vivo could open the way to novel antibiotic treatment strategies. Copyright © 2018 American Society for Microbiology.

Ex vivo 12 h bactericidal activity of oral co-amoxiclav (1.125 g) against beta-lactamase-producing Haemophilus influenzae.

PubMed

Bronner, S; Pompei, D; Elkhaïli, H; Dhoyen, N; Monteil, H; Jehl, F

2001-10-01

The aim of the study was to evaluate the in vitro/ex vivo bactericidal activity of a new coamoxiclav single-dose sachet formulation (1 g amoxicillin + 0.125 g clavulanic acid) against a beta-lactamase-producing strain of Haemophilus influenzae. The evaluation covered the 12 h period after antibiotic administration. Serum specimens from the 12 healthy volunteers included in the pharmacokinetic study were pooled by time point and in equal volumes. Eight of 12 pharmacokinetic sampling time points were included in the study. At time points 0.5, 0.75, 1, 1.5, 2.5, 5, 8 and 12 h post-dosing, the kinetics of bactericidal activity were determined for each of the serial dilutions. Each specimen was serially diluted from 1:2 to 1:256. The index of surviving bacteria (ISB) was subsequently determined for each pharmacokinetic time point. For all the serum samples, bactericidal activity was fast (3-6 h), marked (3-6 log(10) reduction in the initial inoculum) and sustained over the 12 h between-dosing interval. The results obtained also confirmed that the potency of the amoxicillin plus clavulanic acid combination was time dependent against the species under study and that the time interval over which the concentrations were greater than the MIC (t > MIC) was 100% for the strain under study. The data thus generated constitute an interesting prerequisite with a view to using co-amoxiclav 1.125 g in a bd oral regimen.

Whole blood bactericidal activity during treatment of pulmonary tuberculosis.

PubMed

Wallis, Robert S; Vinhas, Solange A; Johnson, John L; Ribeiro, Fabíola C; Palaci, Moisés; Peres, Renata L; Sá, Ricardo T; Dietze, Reynaldo; Chiunda, Allan; Eisenach, Kathleen; Ellner, Jerrold J

2003-01-15

The timely evaluation of new drugs that can be used to shorten tuberculosis (TB) treatment will require surrogate markers for relapse. This study examined bactericidal activity against intracellular Mycobacterium tuberculosis in whole blood culture (whole blood bactericidal activity; WBA) during TB treatment. In the absence of chemotherapy, immune mechanisms in patient blood resulted in bacteriostasis, whereas administration of oral chemotherapy resulted in bacillary killing. Total WBA per dose was greater during the intensive phase of treatment than during the continuation phase (mean, -2.32 vs. -1.67 log(10) cfu-days, respectively; P<.001). Cumulative WBA throughout treatment was greater in subjects whose sputum cultures converted to negative by the eighth week of treatment than in those for whom conversion was delayed (mean, -365 vs. -250 log(10) cfu-days; P=.04) and correlated with the rate of decrease of sputum colony-forming unit counts during the first 4 weeks of treatment (P=.018), both of which are indicative of prognosis. These findings indicate that measurement of WBA may have a role in assessing the sterilizing activity of new anti-TB drugs.

Identification and therapeutic potential of a vitronectin binding region of meningococcal msf.

PubMed

Hill, Darryl J; Griffiths, Natalie J; Borodina, Elena; Andreae, Clio A; Sessions, Richard B; Virji, Mumtaz

2015-01-01

The human pathogen Neisseria meningitides (Nm) attains serum resistance via a number of mechanisms, one of which involves binding to the host complement regulator protein vitronectin. We have shown previously that the Meningococcal surface fibril (Msf), a trimeric autotransporter, binds to the activated form of vitronectin (aVn) to increase Nm survival in human serum. In this study, we aimed to identify the aVn-binding region of Msf to assess its potential as an antigen which can elicit antibodies that block aVn binding and/or possess bactericidal properties. Using several recombinant Msf fragments spanning its surface-exposed region, the smallest aVn-binding recombinants were found to span residues 1-86 and 39-124. The use of further deletion constructs and overlapping recombinant Msf fragments suggested that a region of Msf comprising residues 39-82 may be primarily important for aVn binding and that other regions may also be involved but to a lesser extent. Molecular modelling implicated K66 and K68, conserved in all available Msf sequences, to be involved in the interaction. Recombinant fragments which bound to aVn were able to reduce the survival advantage conveyed by aVn-interaction in serum bactericidal assays. Antibodies raised against one such fragment inhibited aVn binding to Msf. In addition, the antibodies enhanced specific killing of Msf-expressing Nm in a dose-dependent manner. Overall, this study identifies an aVn-binding region of Msf, an adhesin known to impart serum resistance properties to the pathogen; and shows that this region of Msf can elicit antibodies with dual properties which reduce pathogen survival within the host and thus has potential as a vaccine antigen.

Identification and Therapeutic Potential of a Vitronectin Binding Region of Meningococcal Msf

PubMed Central

Hill, Darryl J.; Griffiths, Natalie J.; Borodina, Elena; Andreae, Clio A.; Sessions, Richard B.; Virji, Mumtaz

2015-01-01

The human pathogen Neisseria meningitides (Nm) attains serum resistance via a number of mechanisms, one of which involves binding to the host complement regulator protein vitronectin. We have shown previously that the Meningococcal surface fibril (Msf), a trimeric autotransporter, binds to the activated form of vitronectin (aVn) to increase Nm survival in human serum. In this study, we aimed to identify the aVn-binding region of Msf to assess its potential as an antigen which can elicit antibodies that block aVn binding and/or possess bactericidal properties. Using several recombinant Msf fragments spanning its surface-exposed region, the smallest aVn-binding recombinants were found to span residues 1-86 and 39-124. The use of further deletion constructs and overlapping recombinant Msf fragments suggested that a region of Msf comprising residues 39-82 may be primarily important for aVn binding and that other regions may also be involved but to a lesser extent. Molecular modelling implicated K66 and K68, conserved in all available Msf sequences, to be involved in the interaction. Recombinant fragments which bound to aVn were able to reduce the survival advantage conveyed by aVn-interaction in serum bactericidal assays. Antibodies raised against one such fragment inhibited aVn binding to Msf. In addition, the antibodies enhanced specific killing of Msf-expressing Nm in a dose-dependent manner. Overall, this study identifies an aVn-binding region of Msf, an adhesin known to impart serum resistance properties to the pathogen; and shows that this region of Msf can elicit antibodies with dual properties which reduce pathogen survival within the host and thus has potential as a vaccine antigen. PMID:25826209

Immunogenicity and safety of a novel quadrivalent meningococcal conjugate vaccine (MenACWY-CRM) in healthy Korean adolescents and adults.

PubMed

Lee, Hoan Jong; Chung, Moon-Hyun; Kim, Woo Joo; Hong, Young Jin; Choi, Kyong Min; Lee, Jina; Oh, Chi Eun; Welsch, Jo Anne; Kim, Kyung-Hyo; Hong, Ki Bae; Dagnew, Alemnew F; Bock, Hans; Dull, Peter M; Odrljin, Tatjana

2014-11-01

This phase III placebo-controlled study evaluated the immunogenicity and safety of MenACWY-CRM vaccination in healthy Korean adolescents and adults. Serum bactericidal activity with human complement (hSBA) was measured before and 1 month after vaccination against all four meningococcal serogroups. The IgG concentration specific for serogroup W capsular polysaccharide was measured in a subset of subjects in a post-hoc analysis. Adverse reactions were monitored throughout the study. Four hundred and fifty subjects were randomized 2:1 to receive MenACWY-CRM (N=297) or a saline placebo (N=153). MenACWY-CRM induced a good immune response against all four serogroups, with seroprotection rates (hSBA titers ≥8) of 79%, 99%, 98%, and 94% for serogroups A, C, W, and Y, respectively. Seroresponse rates were high for serogroups A, C, and Y, i.e. 76%, 86%, and 69%, respectively; the rate for serogroup W was 28%. MenACWY-CRM vaccine induced serum bactericidal antibodies against all four serogroups in a majority of subjects regardless of their baseline hSBA titers. MenACWY-CRM was generally well tolerated with most reactions being transient and mild to moderate in severity. Findings of this first study of a quadrivalent meningococcal polysaccharide conjugate vaccine in Korean adults and adolescents demonstrated that a single dose of MenACWY-CRM was well tolerated and immunogenic, as indicated by the percentages of subjects with hSBA titers ≥8 (79%, 99%, 98%, and 94% of subjects) and geometric mean titers (48, 231, 147, and 107) against serogroups A, C, W, and Y, respectively, at 1 month post-vaccination.

Safety and immunogenicity of a novel quadrivalent meningococcal CRM-conjugate vaccine given concomitantly with routine vaccinations in infants.

PubMed

Klein, Nicola P; Reisinger, Keith S; Johnston, William; Odrljin, Tatjana; Gill, Christopher J; Bedell, Lisa; Dull, Peter

2012-01-01

In phase II studies, MenACWY-CRM elicited robust immunologic responses in young infants. We now present results from our pivotal phase III infant immunogenicity/safety study. In this open-label phase III study, we randomized full-term 2-month-old infants to 4 doses of MenACWY-CRM coadministered with routine vaccines at 2, 4, 6, and 12 months of age or with routine vaccines alone. We monitored for local and systemic reactions and serious adverse events among all study participants and evaluated for sufficiency of the immune responses to MenACWY-CRM through serum bactericidal activity assay with human complement. Bactericidal antibodies were present in 94% to 100% of subjects against each of the serogroups in MenACWY-CRM after the 4-dose series and were 67% to 97% after the first 3 doses. Geometric mean titers were higher after the fourth dose of MenACWY-CRM compared with a single dose of MenACWY-CRM at 12 months of age for all serogroups (range of ratios, 4.5-38). Responses to 3 doses of routine vaccines coadministered with MenACWY-CRM were noninferior to routine vaccinations alone, except for small differences in pneumococcal serotype 6B responses after dose 3 but not dose 4 and pertactin after dose 3. Inclusion of MenACWY-CRM did not affect the safety or reactogenicity profiles of the routine infant vaccine series. A 4-dose series of MenACWY-CRM was highly immunogenic and well tolerated in young infants, and it can be coadministered with routine infant vaccines. Substantial immunity was conferred after the first 3 doses administered at 2, 4, and 6 months of age.

Inhibitory role of acyl homoserine lactones in hemolytic activity and viability of Streptococcus pyogenes M6 S165

PubMed Central

Saroj, Sunil D.; Holmer, Linda; Berengueras, Júlia M.; Jonsson, Ann-Beth

2017-01-01

Streptococcus pyogenes an adapted human pathogen asymptomatically colonizes the nasopharynx, among other polymicrobial communities. However, information on the events leading to the colonization and expression of virulence markers subject to interspecies and host-bacteria interactions are limited. The interference of acyl homoserine lactones (AHLs) with the hemolytic activity and viability of S. pyogenes M6 S165 was examined. AHLs, with fatty acid side chains ≥12 carbon atoms, inhibited hemolytic activity by downregulating the expression of the sag operon involved in the production of streptolysin S. Inhibitory AHLs upregulated the expression of transcriptional regulator LuxR. Electrophoretic mobility shift assays revealed the interaction of LuxR with the region upstream of sagA. AHL-mediated bactericidal activity observed at higher concentrations (mM range) was an energy-dependent process, constrained by the requirement of glucose and iron. Ferrichrome transporter FtsABCD facilitated transport of AHLs across the streptococcal membrane. The study demonstrates a previously unreported role for AHLs in S. pyogenes virulence. PMID:28303956

Inhibitory role of acyl homoserine lactones in hemolytic activity and viability of Streptococcus pyogenes M6 S165.

PubMed

Saroj, Sunil D; Holmer, Linda; Berengueras, Júlia M; Jonsson, Ann-Beth

2017-03-17

Streptococcus pyogenes an adapted human pathogen asymptomatically colonizes the nasopharynx, among other polymicrobial communities. However, information on the events leading to the colonization and expression of virulence markers subject to interspecies and host-bacteria interactions are limited. The interference of acyl homoserine lactones (AHLs) with the hemolytic activity and viability of S. pyogenes M6 S165 was examined. AHLs, with fatty acid side chains ≥12 carbon atoms, inhibited hemolytic activity by downregulating the expression of the sag operon involved in the production of streptolysin S. Inhibitory AHLs upregulated the expression of transcriptional regulator LuxR. Electrophoretic mobility shift assays revealed the interaction of LuxR with the region upstream of sagA. AHL-mediated bactericidal activity observed at higher concentrations (mM range) was an energy-dependent process, constrained by the requirement of glucose and iron. Ferrichrome transporter FtsABCD facilitated transport of AHLs across the streptococcal membrane. The study demonstrates a previously unreported role for AHLs in S. pyogenes virulence.

In Vitro and In Vivo studies of monoclonal antibodies with prominent bactericidal activity against Burkholderia pseudomallei and Burkholderia mallei.

PubMed

Zhang, Shimin; Feng, Shaw-Huey; Li, Bingjie; Kim, Hyung-Yong; Rodriguez, Joe; Tsai, Shien; Lo, Shyh-Ching

2011-05-01

Our laboratory has developed more than a hundred mouse monoclonal antibodies (MAbs) against Burkholderia pseudomallei and Burkholderia mallei. These antibodies have been categorized into different groups based on their specificities and the biochemical natures of their target antigens. The current study first examined the bactericidal activities of a number of these MAbs by an in vitro opsonic assay. Then, the in vivo protective efficacy of selected MAbs was evaluated using BALB/c mice challenged intranasally with a lethal dose of the bacteria. The opsonic assay using dimethyl sulfoxide-treated human HL-60 cells as phagocytes revealed that 19 out of 47 tested MAbs (40%) have prominent bactericidal activities against B. pseudomallei and/or B. mallei. Interestingly, all MAbs with strong opsonic activities are those with specificity against either the capsular polysaccharides (PS) or the lipopolysaccharides (LPS) of the bacteria. On the other hand, none of the MAbs reacting to bacterial proteins or glycoproteins showed prominent bactericidal activity. Further study revealed that the antigenic epitopes on either the capsular PS or LPS molecules were readily available for binding in intact bacteria, while the epitopes on proteins/glycoproteins were less accessible to the MAbs. Our in vivo study showed that four MAbs reactive to either the capsular PS or LPS were highly effective in protecting mice against lethal bacterial challenge. The result is compatible with that of our in vitro study. The MAbs with the highest protective efficacy are those reactive to either the capsular PS or LPS of the Burkholderia bacteria.

In Vitro and In Vivo Studies of Monoclonal Antibodies with Prominent Bactericidal Activity against Burkholderia pseudomallei and Burkholderia mallei▿

PubMed Central

Zhang, Shimin; Feng, Shaw-Huey; Li, Bingjie; Kim, Hyung-Yong; Rodriguez, Joe; Tsai, Shien; Lo, Shyh-Ching

2011-01-01

Our laboratory has developed more than a hundred mouse monoclonal antibodies (MAbs) against Burkholderia pseudomallei and Burkholderia mallei. These antibodies have been categorized into different groups based on their specificities and the biochemical natures of their target antigens. The current study first examined the bactericidal activities of a number of these MAbs by an in vitro opsonic assay. Then, the in vivo protective efficacy of selected MAbs was evaluated using BALB/c mice challenged intranasally with a lethal dose of the bacteria. The opsonic assay using dimethyl sulfoxide-treated human HL-60 cells as phagocytes revealed that 19 out of 47 tested MAbs (40%) have prominent bactericidal activities against B. pseudomallei and/or B. mallei. Interestingly, all MAbs with strong opsonic activities are those with specificity against either the capsular polysaccharides (PS) or the lipopolysaccharides (LPS) of the bacteria. On the other hand, none of the MAbs reacting to bacterial proteins or glycoproteins showed prominent bactericidal activity. Further study revealed that the antigenic epitopes on either the capsular PS or LPS molecules were readily available for binding in intact bacteria, while the epitopes on proteins/glycoproteins were less accessible to the MAbs. Our in vivo study showed that four MAbs reactive to either the capsular PS or LPS were highly effective in protecting mice against lethal bacterial challenge. The result is compatible with that of our in vitro study. The MAbs with the highest protective efficacy are those reactive to either the capsular PS or LPS of the Burkholderia bacteria. PMID:21450976

Pharmacological synergism of bee venom and melittin with antibiotics and plant secondary metabolites against multi-drug resistant microbial pathogens.

PubMed

Al-Ani, Issam; Zimmermann, Stefan; Reichling, Jürgen; Wink, Michael

2015-02-15

The goal of this study was to investigate the antimicrobial activity of bee venom and its main component, melittin, alone or in two-drug and three-drug combinations with antibiotics (vancomycin, oxacillin, and amikacin) or antimicrobial plant secondary metabolites (carvacrol, benzyl isothiocyanate, the alkaloids sanguinarine and berberine) against drug-sensitive and antibiotic-resistant microbial pathogens. The secondary metabolites were selected corresponding to the molecular targets to which they are directed, being different from those of melittin and the antibiotics. The minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) were evaluated by the standard broth microdilution method, while synergistic or additive interactions were assessed by checkerboard dilution and time-kill curve assays. Bee venom and melittin exhibited a broad spectrum of antibacterial activity against 51 strains of both Gram-positive and Gram-negative bacteria with strong anti-MRSA and anti-VRE activity (MIC values between 6 and 800 µg/ml). Moreover, bee venom and melittin showed significant antifungal activity (MIC values between 30 and 100 µg/ml). Carvacrol displayed bactericidal activity, while BITC exhibited bacteriostatic activity against all MRSA and VRE strains tested (reference strains and clinical isolates), both compounds showed a remarkable fungicidal activity with minimum fungicidal concentration (MFC) values between 30 and 200 µg/ml. The DNA intercalating alkaloid sanguinarine showed bactericidal activity against MRSA NCTC 10442 (MBC 20 µg/ml), while berberine exhibited bacteriostatic activity against MRSA NCTC 10442 (MIC 40 µg/ml). Checkerboard dilution tests mostly revealed synergism of two-drug combinations against all the tested microorganisms with FIC indexes between 0.24 and 0.50, except for rapidly growing mycobacteria in which combinations exerted an additive effect (FICI = 0.75-1). In time-kill assays all three-drug combinations exhibited a powerful bactericidal synergistic effect against MRSA NCTC 10442, VRE ATCC 51299, and E. coli ATCC 25922 with a reduction of more than 3log10 in the colony count after 24 h. Our findings suggest that bee venom and melittin synergistically enhanced the bactericidal effect of several antimicrobial agents when applied in combination especially when the drugs affect several and differing molecular targets. These results could lead to the development of novel or complementary antibacterial drugs against MDR pathogens. Copyright © 2015 Elsevier GmbH. All rights reserved.

Antibacterial and Antibiofilm Activities of Makaluvamine Analogs

PubMed Central

Nijampatnam, Bhavitavya; Nadkarni, Dwayaja H.; Wu, Hui; Velu, Sadanandan E.

2014-01-01

Streptococcus mutans is a key etiological agent in the formation of dental caries. The major virulence factor is its ability to form biofilms. Inhibition of S. mutans biofilms offers therapeutic prospects for the treatment and the prevention of dental caries. In this study, 14 analogs of makaluvamine, a marine alkaloid, were evaluated for their antibacterial activity against S. mutans and for their ability to inhibit S. mutans biofilm formation. All analogs contained the tricyclic pyrroloiminoquinone core of makaluvamines. The structural variations of the analogs are on the amino substituents at the 7-position of the ring and the inclusion of a tosyl group on the pyrrole ring N of the makaluvamine core. The makaluvamine analogs displayed biofilm inhibition with IC50 values ranging from 0.4 μM to 88 μM. Further, the observed bactericidal activity of the majority of the analogs was found to be consistent with the anti-biofilm activity, leading to the conclusion that the anti-biofilm activity of these analogs stems from their ability to kill S. mutans. However, three of the most potent N-tosyl analogs showed biofilm IC50 values at least an order of magnitude lower than that of bactericidal activity, indicating that the biofilm activity of these analogs is more selective and perhaps independent of bactericidal activity. PMID:25767719

Cross-over assessment of serum bactericidal activity of moxifloxacin and levofloxacin versus penicillin-susceptible and penicillin-resistant Streptococcus pneumoniae in healthy volunteers.

PubMed

Hart, Daniel; Weinstein, Melvin P

2007-07-01

We compared the serum bactericidal activity (SBA) of moxifloxacin and levofloxacin against penicillin-susceptible and penicillin-resistant Streptococcus pneumoniae in 12 healthy volunteers. Each subject received 3 days of oral moxifloxacin 400 mg daily and levofloxacin 750 mg daily, respectively, with a 2- to 4-week washout period between regimens. Blood was drawn at 6 time points after the third dose of each antibiotic. Mean serum bactericidal titers (MSBTRs) for moxifloxacin were 4-fold higher than the mean titers for levofloxacin at each time point. For each drug, MSBTRs at each time point were the same or within one 2-fold dilution when analyzed according to the penicillin susceptibility of the strains or the sex of the subjects. The difference in SBA of the 2 drugs may have implications for the emergence of resistance and clinical outcome.

The Integral Method, a new approach to quantify bactericidal activity.

PubMed

Gottardi, Waldemar; Pfleiderer, Jörg; Nagl, Markus

2015-08-01

The bactericidal activity (BA) of antimicrobial agents is generally derived from the results of killing assays. A reliable quantitative characterization and particularly a comparison of these substances, however, are impossible with this information. We here propose a new method that takes into account the course of the complete killing curve for assaying BA and that allows a clear-cut quantitative comparison of antimicrobial agents with only one number. The new Integral Method, based on the reciprocal area below the killing curve, reliably calculates an average BA [log10 CFU/min] and, by implementation of the agent's concentration C, the average specific bactericidal activity SBA=BA/C [log10 CFU/min/mM]. Based on experimental killing data, the pertaining BA and SBA values of exemplary active halogen compounds were established, allowing quantitative assertions. N-chlorotaurine (NCT), chloramine T (CAT), monochloramine (NH2Cl), and iodine (I2) showed extremely diverging SBA values of 0.0020±0.0005, 1.11±0.15, 3.49±0.22, and 291±137log10 CFU/min/mM, respectively, against Staphylococcus aureus. This immediately demonstrates an approximately 550-fold stronger activity of CAT, 1730-fold of NH2Cl, and 150,000-fold of I2 compared to NCT. The inferred quantitative assertions and conclusions prove the new method suitable for characterizing bactericidal activity. Its application comprises the effect of defined agents on various bacteria, the consequence of temperature shifts, the influence of varying drug structure, dose-effect relationships, ranking of isosteric agents, comparison of competing commercial antimicrobial formulations, and the effect of additives. Copyright © 2015 Elsevier B.V. All rights reserved.

Modulation of post-stroke degenerative and regenerative processes and subacute protection by site-targeted inhibition of the alternative pathway of complement.

PubMed

Alawieh, Ali; Elvington, Andrew; Zhu, Hong; Yu, Jin; Kindy, Mark S; Atkinson, Carl; Tomlinson, Stephen

2015-12-30

Complement promotes neuroinflammation and injury in models of stroke. However, complement is also being increasingly implicated in repair and regeneration after central nervous system (CNS) injury, and some complement deficiencies have been shown to provide acute, but not subacute, protection after murine stroke. Here, we investigate the dual role of complement in injury and repair after cerebral ischemia and reperfusion. We used complement-deficient mice and different complement inhibitors in a model of transient middle cerebral artery occlusion to investigate complement-dependent cellular and molecular changes that occur through the subacute phase after stroke. C3 deficiency and site-targeted complement inhibition with either CR2-Crry (inhibits all pathways) or CR2-fH (inhibits alternative pathway) significantly reduced infarct size, reduced apoptotic cell death, and improved neurological deficit score in the acute phase after stroke. However, only in CR2-fH-treated mice was there sustained protection with no evolution of injury in the subacute phase. Whereas both inhibitors significantly reduced microglia/macrophage activation and astrogliosis in the subacute phase, only CR2-fH improved neurological deficit and locomotor function, maintained neurogenesis markers, enhanced neuronal migration, and increased VEGF expression. These findings in CR2-fH-treated mice correlated with improved performance in spatial learning and passive avoidance tasks. The complement anaphylatoxins have been implicated in repair and regenerative mechanisms after CNS injury, and in this context CR2-fH significantly reduced, but did not eliminate the generation of C5a within the brain, unlike CR2-Crry that completely blocked C5a generation. Gene expression profiling revealed that CR2-fH treatment downregulated genes associated with apoptosis, TGFβ signaling, and neutrophil activation, and decreased neutrophil infiltration was confirmed by immunohistochemistry. CR2-fH upregulated genes for neural growth factor and mediators of neurogenesis and neuronal migration. Live animal imaging demonstrated that following intravenous injection, CR2-fH targeted specifically to the post-ischemic brain, with a tissue half-life of 48.5 h. Finally, unlike C3 deficiency, targeted complement inhibition did not increase susceptibility to lethal post-stroke infection, an important consideration for stroke patients. Ischemic brain tissue-targeted and selective inhibition of alternative complement pathway provide self-limiting inhibition of complement activation and reduces acute injury while maintaining complement-dependent recovery mechanisms into the subacute phase after stroke.

Complement and the control of HIV infection: an evolving story.

PubMed

Frank, Michael M; Hester, Christopher; Jiang, Haixiang

2014-05-01

Thirty years ago, investigators isolated and later determined the structure of HIV-1 and its envelope proteins. Using techniques that were effective with other viruses, they prepared vaccines designed to generate antibody or T-cell responses, but they were ineffective in clinical trials. In this article, we consider the role of complement in host defense against enveloped viruses, the role it might play in the antibody response and why complement has not controlled HIV-1 infection. Complement consists of a large group of cell-bound and plasma proteins that are an integral part of the innate immune system. They provide a first line of defense against microbes and also play a role in the immune response. Here we review the studies of complement-mediated HIV destruction and the role of complement in the HIV antibody response. HIV-1 has evolved a complex defense to prevent complement-mediated killing reviewed here. As part of these studies, we have discovered that HIV-1 envelope, on administration into animals, is rapidly broken down into small peptides that may prove to be very inefficient at provident the type of antigenic stimulation that leads to an effective immune response. Improving complement binding and stabilizing envelope may improve the vaccine response.

Complement therapeutics in inflammatory diseases: promising drug candidates for C3-targeted intervention.

PubMed

Mastellos, D C; Ricklin, D; Hajishengallis, E; Hajishengallis, G; Lambris, J D

2016-02-01

There is increasing appreciation that complement dysregulation lies at the heart of numerous immune-mediated and inflammatory disorders. Complement inhibitors are therefore being evaluated as new therapeutic options in various clinical translation programs and the first clinically approved complement-targeted drugs have profoundly impacted the management of certain complement-mediated diseases. Among the many members of the intricate protein network of complement, the central component C3 represents a 'hot-spot' for complement-targeted therapeutic intervention. C3 modulates both innate and adaptive immune responses and is linked to diverse immunomodulatory systems and biological processes that affect human pathophysiology. Compelling evidence from preclinical disease models has shown that C3 interception may offer multiple benefits over existing therapies or even reveal novel therapeutic avenues in disorders that are not commonly regarded as complement-driven, such as periodontal disease. Using the clinically developed compstatin family of C3 inhibitors and periodontitis as illustrative examples, this review highlights emerging therapeutic concepts and developments in the design of C3-targeted drug candidates as novel immunotherapeutics for oral and systemic inflammatory diseases. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

How Artefacts Mediate Small-Group Co-Creation Activities in a Mobile-Assisted Seamless Language Learning Environment?

ERIC Educational Resources Information Center

Wong, L. -H.; Chen, W.; Jan, M.

2012-01-01

The rich learning resources and contexts learners experience in their everyday life could play important roles in complementing formal learning, but are often neglected by learners and teachers. In this paper, we present an intervention study in "Move, Idioms!", a mobile-assisted Chinese language learning approach that emphasizes contextualized…

In vitro observations on the suitability of new rifamycins for the intermittent chemotherapy of tuberculosis.

PubMed

Dickinson, J M; Mitchison, D A

1987-09-01

The bactericidal activity of six new rifamycin derivatives--rifabutin (RBU), FCE 22250 (F22), rifapentine (RPE), CGP 29861 (C29), CGP 7040 (C70) and CGP 27557 (C27) and rifampicin (RMP)--have been measured against log phase and, as a better test of sterilising activity, against stationary phase cultures of Mycobacterium tuberculosis, H37Rv. The order of activity of 1.0 and 0.2 mg/l rifamycin against log phase cultures was RMP greater than RPE & C27 greater than RBU & C29 greater than C70. The order of activity of 1.0 and 0.4 mg/l, adjusted for stability of the rifamycin, against stationary phase cultures was F22 & RMP greater than RBU greater than RPE greater than C27 & C29 greater than C70. Viable counts were done during and after pulsed exposures of 6, 24 or 96 h to C29 and RMP. The curves were similar though C29 was less bactericidal and the lag period before recovery was 1-2 days longer. F22, having high bactericidal activity against stationary organisms and a long half-life, was considered likely to be the most effective sterilising drug.

Powerful bacterial killing by buckwheat honeys is concentration-dependent, involves complete DNA degradation and requires hydrogen peroxide.

PubMed

Brudzynski, Katrina; Abubaker, Kamal; Wang, Tony

2012-01-01

Exposure of bacterial cells to honey inhibits their growth and may cause cell death. Our previous studies showed a cause-effect relationship between hydroxyl radical generated from honey hydrogen peroxide and growth arrest. Here we explored the role of hydroxyl radicals as inducers of bacterial cells death. The bactericidal effect of ·OH on antibiotic-resistant clinical isolates of MRSA and VRE and standard bacterial strains of E. coli and B. subtiles was examined using a broth microdilution assay supplemented with 3'-(p-aminophenyl) fluorescein (APF) as the ·OH trap, followed by colony enumeration. Bactericidal activities of eight honeys (six varieties of buckwheat, blueberry and manuka honeys) were analyzed. The MBC/MIC ratio ≤4 and the killing curves indicated that honeys exhibited powerful, concentration-dependent bactericidal effect. The extent of killing depended on the ratio of honey concentration to bacterial load, indicating that honey dose was critical for its bactericidal efficacy. The killing rate and potency varied between honeys and ranged from over a 6-log(10) to 4-log(10) CFU/ml reduction of viable cells, equivalent to complete bacterial eradication. The maximal killing was associated with the extensive degradation of bacterial DNA. Honey concentration at which DNA degradation occurred correlated with cell death observed in the concentration-dependent cell-kill on agar plates. There was no quantitative relationship between the ·OH generation by honey and bactericidal effect. At the MBC, where there was no surviving cells and no DNA was visible on agarose gels, the ·OH levels were on average 2-3x lower than at Minimum Inhibitory Concentration (MICs) (p < 0.0001). Pre-treatment of honey with catalase, abolished the bactericidal effect. This raised possibilities that either the abrupt killing prevented accumulation of ·OH (dead cells did not generate ·OH) or that DNA degradation and killing is the actual footprint of ·OH action. In conclusion, honeys of buckwheat origin exhibited powerful, concentration-dependent bactericidal effect. The killing and DNA degradation showed a cause-effect relationship. Hydrogen peroxide was an active part of honey killing mechanism.

Bactericidal activity of cerumen.

PubMed Central

Chai, T J; Chai, T C

1980-01-01

Freshly collected cerumen (dry form) suspended at a concentration of 3% in glycerol-sodium bicarbonate buffer showed bactericidal activity against some strains of bacteria tested. This suspension reduced the viability of Haemophilus influenzae, Escherichia coli K-12, and Serratia marcescens by more than 99%, whereas the viability of two Pseudomonas aeruginosa isolates, E. coli K-1, Streptococcus, and two Staphylococcus aureus isolates of human origin was reduced by 30 to 80%. The results support the hypothesis that cerumen functions to kill certain foreign organisms which enter the ear canal. Images PMID:7447422

In vitro activities of ciprofloxacin and rifampin alone and in combination against growing and nongrowing strains of methicillin-susceptible and methicillin-resistant Staphylococcus aureus.

PubMed Central

Bahl, D; Miller, D A; Leviton, I; Gialanella, P; Wolin, M J; Liu, W; Perkins, R; Miller, M H

1997-01-01

We characterized the effects of ciprofloxacin and rifampin alone and in combination on Staphylococcus aureus in vitro. The effects of drug combinations (e.g., indifferent, antagonistic, or additive interactions) on growth inhibition were compared by disk approximation studies and by determining the fractional inhibitory concentrations. Bactericidal effects in log-phase bacteria and in nongrowing isolates were characterized by time-kill methods. The effect of drug combinations was dependent upon whether or not cells were growing and whether killing or growth inhibition was the endpoint used to measure drug interaction. Despite bactericidal antagonism in time-kill experiments, our in vitro studies suggest several possible explanations for the observed benefits in patients treated with a combination of ciprofloxacin and rifampin for deep-seated staphylococcal infections. Notably, when growth inhibition rather than killing was used to characterize drug interaction, indifference rather than antagonism was observed. An additive bactericidal effect was observed in nongrowing bacteria suspended in phosphate-buffered saline. While rifampin antagonized the bactericidal effects of ciprofloxacin, ciprofloxacin did not antagonize the bactericidal effects of rifampin. Each antimicrobial prevented the emergence of subpopulations that were resistant to the other. PMID:9174186

Antibody-Mediated Complement C3b/iC3b Binding to Group B Streptococcus in Paired Mother and Baby Serum Samples in a Refugee Population on the Thailand-Myanmar Border

PubMed Central

Herbert, Jenny; Thomas, Stephen; Brookes, Charlotte; Turner, Claudia; Turner, Paul; Nosten, Francois; Le Doare, Kirsty; Hudson, Michael; Heath, Paul T.; Gorringe, Andrew

2015-01-01

Streptococcus agalactiae (group B streptococcus [GBS]) is the leading cause of neonatal sepsis and meningitis. In this study, we determined antibody-mediated deposition of complement C3b/iC3b onto the bacterial cell surface of GBS serotypes Ia, Ib, II, III, and V. This was determined for 520 mother and umbilical cord serum sample pairs obtained at the time of birth from a population on the Thailand-Myanmar border. Antibody-mediated deposition of complement C3b/iC3b was detected to at least one serotype in 91% of mothers, despite a known carriage rate in this population of only 12%. Antibody-mediated C3b/iC3b deposition corresponded to known carriage rates, with the highest levels of complement deposition observed onto the most prevalent serotype (serotype II) followed by serotypes Ia, III, V, and Ib. Finally, neonates born to mothers carrying serotype II GBS at the time of birth showed higher antibody-mediated C3b/iC3b deposition against serotype II GBS than neonates born to mothers with no serotype II carriage. Assessment of antibody-mediated C3b/iC3b deposition against GBS may provide insights into the seroepidemiology of anti-GBS antibodies in mothers and infants in different populations. PMID:25589553

Safety and immunogenicity of a booster dose of meningococcal (groups A, C, W, and Y) polysaccharide diphtheria toxoid conjugate vaccine.

PubMed

Robertson, Corwin A; Greenberg, David P; Hedrick, James; Pichichero, Michael; Decker, Michael D; Saunders, Martha

2016-10-17

Quadrivalent meningococcal conjugate vaccines (MenACWY) were developed to offer long-term protection against invasive disease caused by serogroups A, C, W, and Y. Reduced MenACWY effectiveness within 5 years after primary vaccination (likely due to declining bactericidal antibody titers) has been described, particularly with respect to C and Y disease in the United States. We evaluated the safety and immunogenicity of a single booster dose of quadrivalent meningococcal polysaccharide diphtheria toxoid conjugate vaccine (MenACWY-D) in adolescents and adults who received a previous dose 4-6 years earlier. This phase 2, open-label, multicenter study of 834 persons was conducted in the United States. Participants received a single 0.5-mL booster dose of MenACWY-D. Serogroup-specific bactericidal antibody geometric mean titers (GMTs) were measured with a serum bactericidal antibody assay using human complement (hSBA). Proportions of participants achieving antibody titers of ⩾1:8 for each vaccine serogroup on Days 6 and 28 were determined. Rates of adverse events (AEs), including serious adverse events (SAEs), were also assessed. Before booster vaccination, 38.7-68.5% of participants had an hSBA titer ⩾1:8, depending on vaccine serogroup. By Day 6 post-vaccination, 98.2-99.1% of participants had hSBA titers ⩾1:8. By Day 28, >99% of participants achieved this threshold and the primary hypothesis (lower limit of the one-sided 95% confidence limit ⩾85% for each serogroup) was met. The GMT ratios (post-vaccination divided by pre-vaccination) at Day 28 ranged from 47.2 (serogroup A) to 209.1 (serogroup Y). Rates of AEs, including SAEs, were similar to those observed among adolescents and adults who received a primary dose of MenACWY-D in previous studies. There were no study discontinuations due to an AE and no deaths. Booster vaccination with MenACWY-D was safe and induced robust bactericidal antibody responses, consistent with immune memory, among adolescents and adults 4-6 years after primary vaccination. ClinicalTrials.gov registration: NCT01442675. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Antibody ligation of murine Ly-6G induces neutropenia, blood flow cessation, and death via complement-dependent and independent mechanisms.

PubMed

Abbitt, Katherine B; Cotter, Matthew J; Ridger, Victoria C; Crossman, David C; Hellewell, Paul G; Norman, Keith E

2009-01-01

Ly-6G is a member of the Ly-6 family of GPI-linked proteins, which is expressed on murine neutrophils. Antibodies against Ly-6G cause neutropenia, and fatal reactions also develop if mice are primed with TNF-alpha prior to antibody treatment. We have investigated the mechanisms behind these responses to Ly-6G ligation in the belief that similar mechanisms may be involved in neutropenia and respiratory disorders associated with alloantibody ligation of the related Ly-6 family member, NB1, in humans. Neutrophil adhesion, microvascular obstruction, breathing difficulties, and death initiated by anti-Ly-6G antibodies in TNF-alpha-primed mice were shown to be highly complement-dependent, partly mediated by CD11b, CD18, and FcgammaR and associated with clustering of Ly-6G. Neutrophil depletion, on the other hand, was only partly complement-dependent and was not altered by blockade of CD11b, CD18, or FcgammaR. Unlike other neutrophil-activating agents, Ly-6G ligation did not induce neutropenia via sequestration in the lungs. Cross-linking Ly-6G mimicked the responses seen with whole antibody in vivo and also activated murine neutrophils in vitro. Although this suggests that the responses are, in part, mediated by nonspecific properties of antibody ligation, neutrophil depletion requires an additional mechanism possibly specific to the natural function of Ly-6G.

Functional basis for complement evasion by staphylococcal superantigen-like 7.

PubMed

Bestebroer, Jovanka; Aerts, Piet C; Rooijakkers, Suzan H M; Pandey, Manoj K; Köhl, Jörg; van Strijp, Jos A G; de Haas, Carla J C

2010-10-01

The human pathogen Staphylococcus aureus has a plethora of virulence factors that promote its colonization and survival in the host. Among such immune modulators are staphylococcal superantigen-like (SSL) proteins, comprising a family of 14 small, secreted molecules that seem to interfere with the host innate immune system. SSL7 has been described to bind immunoglobulin A (IgA) and complement C5, thereby inhibiting IgA-FcαRI binding and serum killing of Escherichia coli. As C5a generation, in contrast to C5b-9-mediated lysis, is crucial for immune defence against staphylococci, we investigated the impact of SSL7 on staphylococcal-induced C5a-mediated effects. Here, we show that SSL7 inhibits C5a generation induced by staphylococcal opsonization, slightly enhanced by its IgA-binding capacity. Moreover, we demonstrate a strong protective activity of SSL7 against staphylococcal clearance in human whole blood. SSL7 strongly inhibited the C5a-induced phagocytosis of S. aureus and oxidative burst in an in vitro whole-blood inflammation model. Furthermore, we found that SSL7 affects all three pathways of complement activation and inhibits the cleavage of C5 by interference of its binding to C5 convertases. Finally, SSL7 effects were also demonstrated in vivo. In a murine model of immune complex peritonitis, SSL7 abrogated the C5a-driven influx of neutrophils in mouse peritoneum. © 2010 Blackwell Publishing Ltd.

Functional basis for complement evasion by staphylococcal superantigen-like 7

PubMed Central

Bestebroer, Jovanka; Aerts, Piet C.; Rooijakkers, Suzan H.M.; Pandey, Manoj K.; Köhl, Jörg; van Strijp, Jos A. G.; de Haas, Carla J. C.

2010-01-01

Summary The human pathogen Staphylococcus aureus has a plethora of virulence factors that promote its colonization and survival in the host. Among such immune modulators are staphylococcal superantigen-like (SSL) proteins, comprising a family of 14 small, secreted molecules that seem to interfere with the host innate immune system. SSL7 has been described to bind immunoglobulin A (IgA) and complement C5, thereby inhibiting IgA-FcαRI binding and serum killing of E. coli. As C5a generation, in contrast to C5b-9-mediated lysis, is crucial for immune defense against staphylococci, we investigated the impact of SSL7 on staphylococcal-induced C5a-mediated effects. Here, we show that SSL7 inhibits C5a generation induced by staphylococcal opsonization, slightly enhanced by its IgA-binding capacity. Moreover, we demonstrate a strong protective activity of SSL7 against staphylococcal clearance in human whole blood. SSL7 strongly inhibited the C5a-induced phagocytosis of S. aureus and oxidative burst in an in vitro whole blood inflammation model. Furthermore, we found that SSL7 affects all three pathways of complement activation and inhibits the cleavage of C5 by interference of its binding to C5 convertases. Finally, SSL7 effects were also demonstrated in vivo. In a murine model of immune complex peritonitis, SSL7 abrogated the C5a-driven influx of neutrophils in mouse peritoneum. PMID:20545943

Polyanion-Induced Self Association of Complement Factor H1

PubMed Central

Pangburn, Michael K.; Rawal, Nenoo; Cortes, Claudio; Alam, M. Nurul; Ferreira, Viviana P.; Atkinson, Mark A. L.

2008-01-01

Factor H is the primary soluble regulator of activation of the alternative pathway of complement. It prevents activation of complement on host cells and tissues upon association with C3b and surface polyanions such as sialic acids, heparin and other glycosaminoglycans. Here we show that interaction with polyanions causes self-association forming tetramers of the 155,000 Da glycosylated protein. Monomeric human factor H is an extended flexible protein that exhibits an apparent size of 330,000 Da, relative to globular standards, during gel filtration chromatography in the absence of polyanions. In the presence of dextran sulfate (5,000 Da) or heparin an intermediate species of apparent m.w. 700,000 and a limit species of m.w. 1,400,000 were observed by gel filtration. Sedimentation equilibrium analysis by analytical ultracentrifugation indicated a monomer Mr of 163,000 in the absence of polyanions and a Mr of 607,000, corresponding to a tetramer, in the presence of less than a 2-fold molar excess of dextran sulfate. Increasing concentrations of dextran sulfate increased binding of factor H to zymosan-C3b 4.5-fold. This was accompanied by an increase in both the decay accelerating and cofactor activity of factor H on these cells. An expressed fragment encompassing the C-terminal polyanion binding site (complement control protein domains 18–20) also exhibited polyanion-induced self association, suggesting that the C-terminal ends of factor H mediate self-association. The results suggest that recognition of polyanionic markers on host cells and tissues by factor H, and the resulting regulation of complement activation, may involve formation of dimers and tetramers of factor H. PMID:19124749

Loss of Humoral and Cellular Immunity to Invasive Nontyphoidal Salmonella during Current or Convalescent Plasmodium falciparum Infection in Malawian Children

PubMed Central

Nyirenda, James T.; Tembo, Dumizulu L.; Storm, Janet; Dube, Queen; Msefula, Chisomo L.; Jambo, Kondwani C.; Mwandumba, Henry C.; Heyderman, Robert S.; Gordon, Melita A.

2017-01-01

ABSTRACT Invasive nontyphoidal Salmonella (iNTS) infections are commonly associated with Plasmodium falciparum infections, but the immunologic basis for this linkage is poorly understood. We hypothesized that P. falciparum infection compromises the humoral and cellular immunity of the host to NTS, which increases the susceptibility of the host to iNTS infection. We prospectively recruited children aged between 6 and 60 months at a Community Health Centre in Blantyre, Malawi, and allocated them to the following groups; febrile with uncomplicated malaria, febrile malaria negative, and nonfebrile malaria negative. Levels of Salmonella enterica serovar Typhimurium-specific serum bactericidal activity (SBA) and whole-blood bactericidal activity (WBBA), complement C3 deposition, and neutrophil respiratory burst activity (NRBA) were measured. Levels of SBA with respect to S. Typhimurium were reduced in febrile P. falciparum-infected children (median, −0.20 log10 [interquartile range {IQR}, −1.85, 0.32]) compared to nonfebrile malaria-negative children (median, −1.42 log10 [IQR, −2.0, −0.47], P = 0.052). In relation to SBA, C3 deposition on S. Typhimurium was significantly reduced in febrile P. falciparum-infected children (median, 7.5% [IQR, 4.1, 15.0]) compared to nonfebrile malaria-negative children (median, 29% [IQR, 11.8, 48.0], P = 0.048). WBBA with respect to S. Typhimurium was significantly reduced in febrile P. falciparum-infected children (median, 0.25 log10 [IQR, −0.73, 1.13], P = 0.0001) compared to nonfebrile malaria-negative children (median, −1.0 log10 [IQR, −1.68, −0.16]). In relation to WBBA, S. Typhimurium-specific NRBA was reduced in febrile P. falciparum-infected children (median, 8.8% [IQR, 3.7, 20], P = 0.0001) compared to nonfebrile malaria-negative children (median, 40.5% [IQR, 33, 65.8]). P. falciparum infection impairs humoral and cellular immunity to S. Typhimurium in children during malaria episodes, which may explain the increased risk of iNTS observed in children from settings of malaria endemicity. The mechanisms underlying humoral immunity impairment are incompletely understood and should be explored further. PMID:28515136

Loss of Humoral and Cellular Immunity to Invasive Nontyphoidal Salmonella during Current or Convalescent Plasmodium falciparum Infection in Malawian Children.

PubMed

Nyirenda, Tonney S; Nyirenda, James T; Tembo, Dumizulu L; Storm, Janet; Dube, Queen; Msefula, Chisomo L; Jambo, Kondwani C; Mwandumba, Henry C; Heyderman, Robert S; Gordon, Melita A; Mandala, Wilson L

2017-07-01

Invasive nontyphoidal Salmonella (iNTS) infections are commonly associated with Plasmodium falciparum infections, but the immunologic basis for this linkage is poorly understood. We hypothesized that P. falciparum infection compromises the humoral and cellular immunity of the host to NTS, which increases the susceptibility of the host to iNTS infection. We prospectively recruited children aged between 6 and 60 months at a Community Health Centre in Blantyre, Malawi, and allocated them to the following groups; febrile with uncomplicated malaria, febrile malaria negative, and nonfebrile malaria negative. Levels of Salmonella enterica serovar Typhimurium-specific serum bactericidal activity (SBA) and whole-blood bactericidal activity (WBBA), complement C3 deposition, and neutrophil respiratory burst activity (NRBA) were measured. Levels of SBA with respect to S Typhimurium were reduced in febrile P. falciparum -infected children (median, -0.20 log10 [interquartile range {IQR}, -1.85, 0.32]) compared to nonfebrile malaria-negative children (median, -1.42 log10 [IQR, -2.0, -0.47], P = 0.052). In relation to SBA, C3 deposition on S Typhimurium was significantly reduced in febrile P. falciparum -infected children (median, 7.5% [IQR, 4.1, 15.0]) compared to nonfebrile malaria-negative children (median, 29% [IQR, 11.8, 48.0], P = 0.048). WBBA with respect to S Typhimurium was significantly reduced in febrile P. falciparum -infected children (median, 0.25 log10 [IQR, -0.73, 1.13], P = 0.0001) compared to nonfebrile malaria-negative children (median, -1.0 log10 [IQR, -1.68, -0.16]). In relation to WBBA, S Typhimurium-specific NRBA was reduced in febrile P. falciparum -infected children (median, 8.8% [IQR, 3.7, 20], P = 0.0001) compared to nonfebrile malaria-negative children (median, 40.5% [IQR, 33, 65.8]). P. falciparum infection impairs humoral and cellular immunity to S Typhimurium in children during malaria episodes, which may explain the increased risk of iNTS observed in children from settings of malaria endemicity. The mechanisms underlying humoral immunity impairment are incompletely understood and should be explored further. Copyright © 2017 Nyirenda et al.

Differential Mechanism of Escherichia coli Inactivation by (+)-Limonene as a Function of Cell Physiological State and Drug's Concentration

PubMed Central

Chueca, Beatriz; Pagán, Rafael; García-Gonzalo, Diego

2014-01-01

(+)-limonene is a lipophilic antimicrobial compound, extracted from citrus fruits' essential oils, that is used as a flavouring agent and organic solvent by the food industry. A recent study has proposed a common and controversial mechanism of cell death for bactericidal antibiotics, in which hydroxyl radicals ultimately inactivated cells. Our objective was to determine whether the mechanism of Escherichia coli MG1655 inactivation by (+)-limonene follows that of bactericidal antibiotics. A treatment with 2,000 μL/L (+)-limonene inactivated 4 log10 cycles of exponentially growing E. coli cells in 3 hours. On one hand, an increase of cell survival in the ΔacnB mutant (deficient in a TCA cycle enzyme), or in the presence of 2,2′-dipyridyl (inhibitor of Fenton reaction by iron chelation), thiourea, or cysteamine (hydroxyl radical scavengers) was observed. Moreover, the ΔrecA mutant (deficient in an enzyme involved in SOS response to DNA damage) was more sensitive to (+)-limonene. Thus, this indirect evidence indicates that the mechanism of exponentially growing E. coli cells inactivation by 2,000 μL/L (+)-limonene is due to the TCA cycle and Fenton-mediated hydroxyl radical formation that caused oxidative DNA damage, as observed for bactericidal drugs. However, several differences have been observed between the proposed mechanism for bactericidal drugs and for (+)-limonene. In this regard, our results demonstrated that E. coli inactivation was influenced by its physiological state and the drug's concentration: experiments with stationary-phase cells or 4,000 μL/L (+)-limonene uncovered a different mechanism of cell death, likely unrelated to hydroxyl radicals. Our research has also shown that drug's concentration is an important factor influencing the mechanism of bacterial inactivation by antibiotics, such as kanamycin. These results might help in improving and spreading the use of (+)-limonene as an antimicrobial compound, and in clarifying the controversy about the mechanism of inactivation by bactericidal antibiotics. PMID:24705541

Differential mechanism of Escherichia coli Inactivation by (+)-limonene as a function of cell physiological state and drug's concentration.

PubMed

Chueca, Beatriz; Pagán, Rafael; García-Gonzalo, Diego

2014-01-01

(+)-limonene is a lipophilic antimicrobial compound, extracted from citrus fruits' essential oils, that is used as a flavouring agent and organic solvent by the food industry. A recent study has proposed a common and controversial mechanism of cell death for bactericidal antibiotics, in which hydroxyl radicals ultimately inactivated cells. Our objective was to determine whether the mechanism of Escherichia coli MG1655 inactivation by (+)-limonene follows that of bactericidal antibiotics. A treatment with 2,000 μL/L (+)-limonene inactivated 4 log10 cycles of exponentially growing E. coli cells in 3 hours. On one hand, an increase of cell survival in the ΔacnB mutant (deficient in a TCA cycle enzyme), or in the presence of 2,2'-dipyridyl (inhibitor of Fenton reaction by iron chelation), thiourea, or cysteamine (hydroxyl radical scavengers) was observed. Moreover, the ΔrecA mutant (deficient in an enzyme involved in SOS response to DNA damage) was more sensitive to (+)-limonene. Thus, this indirect evidence indicates that the mechanism of exponentially growing E. coli cells inactivation by 2,000 μL/L (+)-limonene is due to the TCA cycle and Fenton-mediated hydroxyl radical formation that caused oxidative DNA damage, as observed for bactericidal drugs. However, several differences have been observed between the proposed mechanism for bactericidal drugs and for (+)-limonene. In this regard, our results demonstrated that E. coli inactivation was influenced by its physiological state and the drug's concentration: experiments with stationary-phase cells or 4,000 μL/L (+)-limonene uncovered a different mechanism of cell death, likely unrelated to hydroxyl radicals. Our research has also shown that drug's concentration is an important factor influencing the mechanism of bacterial inactivation by antibiotics, such as kanamycin. These results might help in improving and spreading the use of (+)-limonene as an antimicrobial compound, and in clarifying the controversy about the mechanism of inactivation by bactericidal antibiotics.

Scabies Mite Peritrophins Are Potential Targets of Human Host Innate Immunity

PubMed Central

Holt, Deborah C.; Kemp, Dave J.; Fischer, Katja

2011-01-01

Background Pruritic scabies lesions caused by Sarcoptes scabiei burrowing in the stratum corneum of human skin facilitate opportunistic bacterial infections. Emerging resistance to current therapeutics emphasizes the need to identify novel targets for protective intervention. We have characterized several protein families located in the mite gut as crucial factors for host-parasite interactions. Among these multiple proteins inhibit human complement, presumably to avoid complement-mediated damage of gut epithelial cells. Peritrophins are major components of the peritrophic matrix often found in the gut of arthropods. We hypothesized that a peritrophin, if abundant in the scabies mite gut, could be an activator of complement. Methodology/Principal Findings A novel full length scabies mite peritrophin (SsPTP1) was identified in a cDNA library from scabies mites. The amino acid sequence revealed four putative chitin binding domains (CBD). Recombinant expression of one CBD of the highly repetitive SsPTP1 sequence as TSP-hexaHis-fusion protein resulted in soluble protein, which demonstrated chitin binding activity in affinity chromatography assays. Antibodies against a recombinant SsPTP1 fragment were used to immunohistochemically localize native SsPTP1 in the mite gut and in fecal pellets within the upper epidermis, co-localizing with serum components such as host IgG and complement. Enzymatic deglycosylation confirmed strong N- and O-glycosylation of the native peritrophin. Serum incubation followed by immunoblotting with a monoclonal antibody against mannan binding lectin (MBL), the recognition molecule of the lectin pathway of human complement activation, indicated that MBL may specifically bind to glycosylated SsPTP1. Conclusions/Significance This study adds a new aspect to the accumulating evidence that complement plays a major role in scabies mite biology. It identifies a novel peritrophin localized in the mite gut as a potential target of the lectin pathway of the complement cascade. These initial findings indicate a novel role of scabies mite peritrophins in triggering a host innate immune response within the mite gut. PMID:21980545

A novel factor H-Fc chimeric immunotherapeutic molecule against Neisseria gonorrhoeae

PubMed Central

Shaughnessy, Jutamas; Gulati, Sunita; Agarwal, Sarika; Unemo, Magnus; Ohnishi, Makoto; Su, Xia-Hong; Monks, Brian G.; Visintin, Alberto; Madico, Guillermo; Lewis, Lisa A.; Golenbock, Douglas T.; Reed, George W.; Rice, Peter A.; Ram, Sanjay

2015-01-01

Neisseria gonorrhoeae (Ng), the causative agent of the sexually transmitted infection gonorrhea, has developed resistance to almost every conventional antibiotic. There is an urgent need to develop novel therapies against gonorrhea. Many pathogens, including Ng, bind the complement inhibitor factor H (FH) to evade complement-dependent killing. Sialylation of gonococcal lipooligosaccharide, as occurs in vivo, augments binding of human FH through its domains 18-20 (FH18-20). We explored the utility of fusing FH18-20 with IgG Fc (FH18-20/Fc) to create a novel anti-infective immunotherapeutic. FH18-20 also binds to select host glycosaminoglycans to limit unwanted complement activation on host cells. To identify mutation(s) in FH18-20 that eliminated complement activation on host cells, yet maintained binding to Ng, we created four mutations in domains 19 or 20 described in atypical hemolytic uremic syndrome that prevented binding of mutated fH to human erythrocytes. One of the mutant proteins (D to G at position 1119 in domain 19; FHD1119G/Fc) facilitated complement-dependent killing of gonococci similar to unmodified FH18-20/Fc, but unlike FH18-20/Fc, did not lyse human erythrocytes. FHD1119G/Fc bound to all (100%) of 15 sialylated clinical Ng isolates tested (including three contemporary ceftriaxone-resistant strains), mediated complement-dependent killing of 10/15 (67%) strains and enhanced C3 deposition (≥10-fold above baseline levels) on each of the five isolates not directly killed by complement. FHD1119G/Fc facilitated opsonophagocytic killing of a serum-resistant strain by human polymorphonuclear neutrophils. FHD1119G/Fc administered intravaginally significantly reduced the duration and burden of gonococcal infection in the mouse vaginal colonization model. FHD1119G/Fc represents a novel immunotherapeutic against multidrug-resistant Ng. PMID:26773149

Comparative antimicrobial activity, in vitro and in vivo, of soft N-chloramine systems and chlorhexidine.

PubMed Central

Selk, S H; Pogány, S A; Higuchi, T

1982-01-01

Antimicrobial activity of the following four new N-chloramine compounds was evaluated: two chlorinated simple amino acids, a chlorinated half-ester of succinic acid, and a chlorinated half-ester of glutaric acid. For comparison, the known bactericidal agents 3-chloro-4,4-dimethyl-2-oxazolidinone and chlorhexidine were evaluated by the same procedure. The contact germicidal efficiency screen was used to examine the in vitro bactericidal activity of all six compounds in the absence and presence of 5% horse serum or 5% Triton X-100. The four new compounds were found to have greater germicidal activity than the other compounds tested and to exhibit low toxicity and skin irritation values. The in vivo bactericidal activity was evaluated in two studies. In the occlusion test, three of the four new compounds plus chlorhexidine diacetate were tested. The N-chloramines were significantly superior to chlorhexidine in preventing the expansion of the normal flora under occlusion. In the scrub test, a gloved-hand wash method was used to compare the antimicrobial effect of a 1% solution of the chlorinated half-ester of succinic acid in triacetin with that of a commercial germicidal hand wash containing 4% chlorhexidine gluconate. The two preparations exhibited essentially the same hand-degerming activity. PMID:6805433

Soya bean tempe extracts show antibacterial activity against Bacillus cereus cells and spores.

PubMed

Roubos-van den Hil, P J; Dalmas, E; Nout, M J R; Abee, T

2010-07-01

Tempe, a Rhizopus ssp.-fermented soya bean food product, was investigated for bacteriostatic and/or bactericidal effects against cells and spores of the food-borne pathogen Bacillus cereus. Tempe extract showed a high antibacterial activity against B. cereus ATCC 14579 based on optical density and viable count measurements. This growth inhibition was manifested by a 4 log CFU ml(-1) reduction, within the first 15 min of exposure. Tempe extracts also rapidly inactivated B. cereus spores upon germination. Viability and membrane permeability assessments using fluorescence probes showed rapid inactivation and permeabilization of the cytoplasmic membrane confirming the bactericidal mode of action. Cooked beans and Rhizopus grown on different media did not show antibacterial activity, indicating the unique association of the antibacterial activity with tempe. Subsequent characterization of the antibacterial activity revealed that heat treatment and protease addition nullified the bactericidal effect, indicating the proteinaceous nature of the bioactive compound. During fermentation of soya beans with Rhizopus, compounds are released with extensive antibacterial activity against B. cereus cells and spores. The results show the potential of producing natural antibacterial compounds that could be used as ingredients in food preservation and pathogen control. © 2009 The Authors. Journal compilation © 2009 The Society for Applied Microbiology.

Biofilm-forming activity of bacteria isolated from toilet bowl biofilms and the bactericidal activity of disinfectants against the isolates.

PubMed

Mori, Miho; Gomi, Mitsuhiro; Matsumune, Norihiko; Niizeki, Kazuma; Sakagami, Yoshikazu

2013-01-01

To evaluate the sanitary conditions of toilets, the bacterial counts of the toilet bowl biofilms in 5 Kansai area and 11 Kansai and Kanto area homes in Japan were measured in winter and summer seasons, respectively. Isolates (128 strains) were identified by analyzing 16S ribosomal RNA sequences. The number of colonies and bacterial species from biofilms sampled in winter tended to be higher and lower, respectively, than those in summer. Moreover, the composition of bacterial communities in summer and winter samples differed considerably. In summer samples, biofilms in Kansai and Kanto areas were dominated by Blastomonas sp. and Mycobacterium sp., respectively. Methylobacterium sp. was detected in all toilet bowl biofilms except for one sample. Methylobacterium sp. constituted the major presence in biofilms along with Brevundimonas sp., Sphingomonas sp., and/or Pseudomonas sp. The composition ratio of the sum of their genera was 88.0 from 42.9% of the total bacterial flora. The biofilm formation abilities of 128 isolates were investigated, and results suggested that Methylobacterium sp. and Sphingomonas sp. were involved in biofilm formation in toilet bowls. The biofilm formation of a mixed bacteria system that included bacteria with the highest biofilm-forming ability in a winter sample was greater than mixture without such bacteria. This result suggests that isolates possessing a high biofilm-forming activity are involved in the biofilm formation in the actual toilet bowl. A bactericidal test against 25 strains indicated that the bactericidal activities of didecyldimethylammonium chloride (DDAC) tended to be higher than those of polyhexamethylene biguanide (PHMB) and N-benzyl-N,N-dimethyldodecylammonium chloride (ADBAC). In particular, DDAC showed high bactericidal activity against approximately 90% of tested strains under the 5 h treatment.

A novel chimeric phage lysin with high in vitro and in vivo bactericidal activity against Streptococcus pneumoniae.

PubMed

Díez-Martínez, Roberto; De Paz, Héctor D; García-Fernández, Esther; Bustamante, Noemí; Euler, Chad W; Fischetti, Vincent A; Menendez, Margarita; García, Pedro

2015-01-01

Streptococcus pneumoniae is becoming increasingly antibiotic resistant worldwide and new antimicrobials are urgently needed. Our aim was new chimeric phage endolysins, or lysins, with improved bactericidal activity by swapping the structural components of two pneumococcal phage lysozymes: Cpl-1 (the best lysin tested to date) and Cpl-7S. The bactericidal effects of four new chimeric lysins were checked against several bacteria. The purified enzymes were added at different concentrations to resuspended bacteria and viable cells were measured after 1 h. Killing capacity of the most active lysin, Cpl-711, was tested in a mouse bacteraemia model, following mouse survival after injecting different amounts (25-500 μg) of enzyme. The capacity of Cpl-711 to reduce pneumococcal biofilm formation was also studied. The chimera Cpl-711 substantially improved the killing activity of the parental phage lysozymes, Cpl-1 and Cpl-7S, against pneumococcal bacteria, including multiresistant strains. Specifically, 5 μg/mL Cpl-711 killed ≥7.5 log of pneumococcal R6 strain. Cpl-711 also reduced pneumococcal biofilm formation and killed 4 log of the bacterial population at 1 μg/mL. Mice challenged intraperitoneally with D39_IU pneumococcal strain were protected by treatment with a single intraperitoneal injection of Cpl-711 1 h later, resulting in about 50% greater protection than with Cpl-1. Domain swapping among phage lysins allows the construction of new chimeric enzymes with high bactericidal activity and a different substrate range. Cpl-711, the most powerful endolysin against pneumococci, offers a promising therapeutic perspective for the treatment of multiresistant pneumococcal infections. © The Author 2015. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Morphological bactericidal fast-acting effects of peracetic acid, a high-level disinfectant, against Staphylococcus aureus and Pseudomonas aeruginosa biofilms in tubing.

PubMed

Chino, T; Nukui, Y; Morishita, Y; Moriya, K

2017-01-01

The bactericidal effect of disinfectants against biofilms is essential to reduce potential endoscopy-related infections caused by contamination. Here, we investigated the bactericidal effect of a high-level disinfectant, peracetic acid (PAA), against Staphylococcus aureus and Pseudomonas aeruginosa biofilm models in vitro. S. aureus and P. aeruginosa biofilms were cultured at 35 °C for 7 days with catheter tubes. The following high-level disinfectants (HLDs) were tested: 0.3% PAA, 0.55% ortho-phthalaldehyde (OPA), and 2.0% alkaline-buffered glutaraldehyde (GA). Biofilms were exposed to these agents for 1-60 min and observed after 5 min and 30 min by transmission and scanning electron microscopy. A Student's t test was performed to compare the exposure time required for bactericidal effectiveness of the disinfectants. PAA and GA were active within 1 min and 5 min, respectively, against S. aureus and P. aeruginosa biofilms. OPA took longer than 10 min and 30 min to act against S. aureus and P. aeruginosa biofilms, respectively ( p  < 0.01). Treatment with PAA elicited changes in cell shape after 5 min and structural damage after 30 min. Amongst the HLDs investigated, PAA elicited the most rapid bactericidal effects against both biofilms. Additionally, treatment with PAA induced morphological alterations in the in vitro biofilm models, suggesting that PAA exerts fast-acting bactericidal effects against biofilms associated with endoscopy-related infections. These findings indicate that the exposure time for bactericidal effectiveness of HLDs for endoscope reprocessing in healthcare settings should be reconsidered.

A simple and convenient microtiter plate assay for the detection of bactericidal antibodies to Vibrio cholerae O1 and Vibrio cholerae O139.

PubMed

Boutonnier, Alain; Dassy, Bruno; Duménil, Rémy; Guénolé, Alain; Ratsitorahina, Maherisoa; Migliani, René; Fournier, Jean-Michel

2003-12-01

It is believed that the correlate of protection for cholera can be determined by the serum vibriocidal assay. The currently available vibriocidal assays, based on the conventional agar plating technique, are labor intensive. We developed a simple and convenient microtiter plate assay for the detection of vibriocidal antibodies that is equally as efficient for Vibrio cholerae O1 and for V. cholerae O139. The addition of succinate and neotetrazolium made it possible to measure the growth of surviving bacterial target cells by monitoring a color change. We evaluated assay parameters (target strains, growth of target cells, complement source and concentration) that may affect the reproducibility of the method for V. cholerae O139. The results obtained with the microtiter plate assay were uniformly similar to those obtained with the conventional agar plating assay, when testing both the Inaba and Ogawa serotypes of V. cholerae O1. The microtiter plate assay was also convenient for measuring the activity of animal sera and mouse monoclonal antibodies.

Antistaphylococcal activity of DX-619, a new des-F(6)-quinolone, compared to those of other agents.

PubMed

Bogdanovich, Tatiana; Esel, Duygu; Kelly, Linda M; Bozdogan, Bülent; Credito, Kim; Lin, Gengrong; Smith, Kathy; Ednie, Lois M; Hoellman, Dianne B; Appelbaum, Peter C

2005-08-01

The in vitro activity of DX-619, a new des-F(6)-quinolone, was tested against staphylococci and compared to those of other antimicrobials. DX-619 had the lowest MIC ranges/MIC(50)s/MIC(90)s (microg/ml) against 131 Staphylococcus aureus strains (32), and ciprofloxacin (>32/>32). Raised quinolone MICs were associated with mutations in GyrA (S84L) and single or double mutations in GrlA (S80F or Y; E84K, G, or V) in all S. aureus strains tested. A recent vancomycin-resistant S. aureus (VRSA) strain (Hershey) was resistant to available quinolones and was inhibited by DX-619 at 0.25 microg/ml and sitafloxacin at 1.0 microg/ml. Vancomycin (except VRSA), linezolid, ranbezolid, tigecycline, and quinupristin-dalfopristin were active against all strains, and teicoplanin was active against S. aureus but less active against coagulase-negative staphylococci. DX-619 produced resistant mutants with MICs of 1 to >32 microg/ml after <50 days of selection compared to 16 to >32 microg/ml for ciprofloxacin, sitafloxacin, moxifloxacin, and gatifloxacin. DX-619 and sitafloxacin were also more active than other tested drugs against selected mutants and had the lowest mutation frequencies in single-step resistance selection. DX-619 and sitafloxacin were bactericidal against six quinolone-resistant (including the VRSA) and seven quinolone-susceptible strains tested, whereas gatifloxacin, moxifloxacin, levofloxacin, and ciprofloxacin were bactericidal against 11, 10, 7, and 5 strains at 4x MIC after 24 h, respectively. DX-619 was also bactericidal against one other VRSA strain, five vancomycin-intermediate S. aureus strains, and four vancomycin-intermediate coagulase-negative staphylococci. Linezolid, ranbezolid, and tigecycline were bacteriostatic and quinupristin-dalfopristin, teicoplanin, and vancomycin were bactericidal against two, eight, and nine strains, and daptomycin and oritavancin were rapidly bactericidal against all strains, including the VRSA. DX-619 has potent in vitro activity against staphylococci, including methicillin-, ciprofloxacin-, and vancomycin-resistant strains.

Comparative in vitro susceptibilities and bactericidal activities of investigational fluoroquinolone ABT-492 and other antimicrobial agents against human mycoplasmas and ureaplasmas.

PubMed

Waites, Ken B; Crabb, Donna M; Duffy, Lynn B

2003-12-01

We determined in vitro susceptibilities for ABT-492 and other antimicrobials against Mycoplasma pneumoniae, Mycoplasma fermentans, Mycoplasma hominis, and Ureaplasma species. ABT-492 MICs were < or =1 microg/ml, and the agent was bactericidal against selected isolates of M. pneumoniae and M. hominis. ABT-492 has potential for treatment of infections due to these microorganisms.

Comparative In Vitro Susceptibilities and Bactericidal Activities of Investigational Fluoroquinolone ABT-492 and Other Antimicrobial Agents against Human Mycoplasmas and Ureaplasmas

PubMed Central

Waites, Ken B.; Crabb, Donna M.; Duffy, Lynn B.

2003-01-01

We determined in vitro susceptibilities for ABT-492 and other antimicrobials against Mycoplasma pneumoniae, Mycoplasma fermentans, Mycoplasma hominis, and Ureaplasma species. ABT-492 MICs were ≤1 μg/ml, and the agent was bactericidal against selected isolates of M. pneumoniae and M. hominis. ABT-492 has potential for treatment of infections due to these microorganisms. PMID:14638513

Bactericidal activities of woven cotton and nonwoven polypropylene fabrics coated with hydroxyapatite-binding silver/titanium dioxide ceramic nanocomposite “Earth-plus”

PubMed Central

Kasuga, Eriko; Kawakami, Yoshiyuki; Matsumoto, Takehisa; Hidaka, Eiko; Oana, Kozue; Ogiwara, Naoko; Yamaki, Dai; Sakurada, Tsukasa; Honda, Takayuki

2011-01-01

Background Bacteria from the hospital environment, including linens and curtains, are often responsible for hospital-associated infections. The aim of the present study was to evaluate the bactericidal effects of fabrics coated with the hydroxyapatite-binding silver/titanium dioxide ceramic nanocomposite “Earth-plus”. Methods Bactericidal activities of woven and nonwoven fabrics coated with Earth-plus were investigated by the time-kill curve method using nine bacterial strains, including three Staphylococcus aureus, three Escherichia coli, and three Pseudomonas aeruginosa strains. Results The numbers of viable S. aureus and E. coli cells on both fabrics coated with Earth-plus decreased to below 2 log10 colony-forming units/mL in six hours and reached the detection limit in 18 hours. Viable cell counts of P. aeruginosa on both fabrics coated with Earth-plus could not be detected after 3–6 hours. Viable cells on woven fabrics showed a more rapid decline than those on nonwoven fabrics. Bacterial cell counts of the nine strains on fabrics without Earth-plus failed to decrease even after 18 hours. Conclusion Woven cotton and nonwoven polypropylene fabrics were shown to have excellent antibacterial potential. The woven fabric was more bactericidal than the nonwoven fabric. PMID:21931489

Green synthesis and characterization of Carica papaya leaf extract coated silver nanoparticles through X-ray diffraction, electron microscopy and evaluation of bactericidal properties

PubMed Central

Banala, Rajkiran Reddy; Nagati, Veera Babu; Karnati, Pratap Reddy

2015-01-01

The evolution of nanotechnology and the production of nanomedicine from various sources had proven to be of intense value in the field of biomedicine. The smaller size of nanoparticles is gaining importance in research for the treatment of various diseases. Moreover the production of nanoparticles is eco-friendly and cost effective. In the present study silver nanoparticles were synthesized from Carica papaya leaf extract (CPL) and characterized for their size and shape using scanning electron microscopy and transmission electron microscopy, respectively. Fourier transform infrared spectroscopy (FTIR), Energy dispersive X-ray spectroscopy (EDS/EDX) and X-ray diffraction spectroscopy (XRD) were conducted to determine the concentration of metal ions, the shape of molecules. The bactericidal activity was evaluated using Luria Bertani broth cultures and the minimum inhibition concentration (MIC) and minimum bactericidal concentration (MBC) were estimated using turbidimetry. The data analysis showed size of 50–250 nm spherical shaped nanoparticles. The turbidimetry analysis showed MIC and MBC was >25 μg/mL against both Gram positive and Gram negative bacteria in Luria Bertani broth cultures. In summary the synthesized silver nanoparticles from CPL showed acceptable size and shape of nanoparticles and effective bactericidal activity. PMID:26288570

Green synthesis and characterization of Carica papaya leaf extract coated silver nanoparticles through X-ray diffraction, electron microscopy and evaluation of bactericidal properties.

PubMed

Banala, Rajkiran Reddy; Nagati, Veera Babu; Karnati, Pratap Reddy

2015-09-01

The evolution of nanotechnology and the production of nanomedicine from various sources had proven to be of intense value in the field of biomedicine. The smaller size of nanoparticles is gaining importance in research for the treatment of various diseases. Moreover the production of nanoparticles is eco-friendly and cost effective. In the present study silver nanoparticles were synthesized from Carica papaya leaf extract (CPL) and characterized for their size and shape using scanning electron microscopy and transmission electron microscopy, respectively. Fourier transform infrared spectroscopy (FTIR), Energy dispersive X-ray spectroscopy (EDS/EDX) and X-ray diffraction spectroscopy (XRD) were conducted to determine the concentration of metal ions, the shape of molecules. The bactericidal activity was evaluated using Luria Bertani broth cultures and the minimum inhibition concentration (MIC) and minimum bactericidal concentration (MBC) were estimated using turbidimetry. The data analysis showed size of 50-250 nm spherical shaped nanoparticles. The turbidimetry analysis showed MIC and MBC was >25 μg/mL against both Gram positive and Gram negative bacteria in Luria Bertani broth cultures. In summary the synthesized silver nanoparticles from CPL showed acceptable size and shape of nanoparticles and effective bactericidal activity.

Structural basis for complement evasion by Lyme disease pathogen Borrelia burgdorferi.

PubMed

Bhattacharjee, Arnab; Oeemig, Jesper S; Kolodziejczyk, Robert; Meri, Taru; Kajander, Tommi; Lehtinen, Markus J; Iwaï, Hideo; Jokiranta, T Sakari; Goldman, Adrian

2013-06-28

Borrelia burgdorferi spirochetes that cause Lyme borreliosis survive for a long time in human serum because they successfully evade the complement system, an important arm of innate immunity. The outer surface protein E (OspE) of B. burgdorferi is needed for this because it recruits complement regulator factor H (FH) onto the bacterial surface to evade complement-mediated cell lysis. To understand this process at the molecular level, we used a structural approach. First, we solved the solution structure of OspE by NMR, revealing a fold that has not been seen before in proteins involved in complement regulation. Next, we solved the x-ray structure of the complex between OspE and the FH C-terminal domains 19 and 20 (FH19-20) at 2.83 Å resolution. The structure shows that OspE binds FH19-20 in a way similar to, but not identical with, that used by endothelial cells to bind FH via glycosaminoglycans. The observed interaction of OspE with FH19-20 allows the full function of FH in down-regulation of complement activation on the bacteria. This reveals the molecular basis for how B. burgdorferi evades innate immunity and suggests how OspE could be used as a potential vaccine antigen.

Influence of Scaffold Size on Bactericidal Activity of Nitric Oxide Releasing Silica Nanoparticles

PubMed Central

Carpenter, Alexis W.; Slomberg, Danielle L.; Rao, Kavitha S.; Schoenfisch, Mark H.

2011-01-01

A reverse microemulsion synthesis was used to prepare amine functionalized silica nanoparticles of three distinct sizes (i.e., 50, 100, and 200 nm) with identical amine concentrations. The resulting hybrid nanoparticles, consisting of N-(6 aminohexyl) aminopropyltrimethoxysilane and tetraethoxysilane, were highly monodisperse in size. N-diazeniumdiolate nitric oxide (NO) donors were subsequently formed on secondary amines while controlling reaction conditions to keep the total amount of nitric oxide (NO) released constant for each particle size. The bactericidal efficacy of the NO releasing nanoparticles against Pseudomonas aeruginosa increased with decreasing particle size. Additionally, smaller diameter nanoparticles were found to associate with the bacteria at a faster rate and to a greater extent than larger particles. Neither control (non-NO-releasing) nor NO releasing particles exhibited toxicity towards L929 mouse fibroblasts at concentrations above their respective minimum bactericidal concentrations. This study represents the first investigation of the bactericidal efficacy of NO-releasing silica nanoparticles as a function of particle size. PMID:21842899

An enhancer peptide for membrane-disrupting antimicrobial peptides

PubMed Central

2010-01-01

Background NP4P is a synthetic peptide derived from a natural, non-antimicrobial peptide fragment (pro-region of nematode cecropin P4) by substitution of all acidic amino acid residues with amides (i.e., Glu → Gln, and Asp → Asn). Results In the presence of NP4P, some membrane-disrupting antimicrobial peptides (ASABF-α, polymyxin B, and nisin) killed microbes at lower concentration (e.g., 10 times lower minimum bactericidal concentration for ASABF-α against Staphylococcus aureus), whereas NP4P itself was not bactericidal and did not interfere with bacterial growth at ≤ 300 μg/mL. In contrast, the activities of antimicrobial agents with a distinct mode of action (indolicidin, ampicillin, kanamycin, and enrofloxacin) were unaffected. Although the membrane-disrupting activity of NP4P was slight or undetectable, ASABF-α permeabilized S. aureus membranes with enhanced efficacy in the presence of NP4P. Conclusions NP4P selectively enhanced the bactericidal activities of membrane-disrupting antimicrobial peptides by increasing the efficacy of membrane disruption against the cytoplasmic membrane. PMID:20152058

Bactericidal Effects of HVOF-Sprayed Nanostructured TiO2 on Pseudomonas aeruginosa

NASA Astrophysics Data System (ADS)

Jeffery, B.; Peppler, M.; Lima, R. S.; McDonald, A.

2010-01-01

Titanium dioxide (TiO2) has been shown to exhibit photocatalytic bactericidal activity. This preliminary study focused on examining the photocatalytic activity of high-velocity oxy-fuel (HVOF) sprayed nanostructured TiO2 coatings to kill Pseudomonas aeruginosa. The surfaces of the nanostructured TiO2 coatings were lightly polished before addition of the bacterial solution. Plates of P. aeruginosa were grown, and then suspended in a phosphate buffer saline (PBS) solution. The concentration of bacteria used was determined by a photo-spectrometer, which measured the amount of light absorbed by the bacteria-filled solution. This solution was diluted and pipetted onto the coating, which was exposed to white light in 30-min intervals, up to 120 min. It was found that on polished HVOF-sprayed coatings exposed to white light, 24% of the bacteria were killed after exposure for 120 min. On stainless steel controls, approximately 6% of the bacteria were not recovered. These preliminary results show that thermal-sprayed nanostructured TiO2 coatings exhibited photocatalytic bactericidal activity with P. aeruginosa.

Naturally Acquired Antibodies against Haemophilus influenzae Type a in Aboriginal Adults, Canada

PubMed Central

Nix, Eli B.; Williams, Kylie; Cox, Andrew D.; St. Michael, Frank; Romero-Steiner, Sandra; Schmidt, Daniel S.; McCready, William G.

2015-01-01

In the post-Haemophilus influenzae type b (Hib) vaccine era that began in the 1980's, H. influenzae type a (Hia) emerged as a prominent cause of invasive disease in North American Aboriginal populations. To test whether a lack of naturally acquired antibodies may underlie increased rates of invasive Hia disease, we compared serum bactericidal activity against Hia and Hib and IgG and IgM against capsular polysaccharide between Canadian Aboriginal and non-Aboriginal healthy and immunocompromised adults. Both healthy and immunocompromised Aboriginal adults exhibited significantly higher bactericidal antibody titers against Hia than did non-Aboriginal adults (p = 0.042 and 0.045 respectively), with no difference in functional antibody activity against Hib. IgM concentrations against Hia were higher than IgG in most study groups; the inverse was true for antibody concentrations against Hib. Our results indicate that Aboriginal adults possess substantial serum bactericidal activity against Hia that is mostly due to IgM antibodies. The presence of sustained IgM against Hia suggests recent Hia exposure. PMID:25626129

Bactericidal Activity of Micromolar N-Chlorotaurine: Evidence for Its Antimicrobial Function in the Human Defense System

PubMed Central

Nagl, Markus; Hess, Michael W.; Pfaller, Kristian; Hengster, Paul; Gottardi, Waldemar

2000-01-01

N-Chlorotaurine, the main representative of long-lived oxidants found in the supernatant of stimulated granulocytes, has been investigated systematically with regard to its antibacterial activity at different physiological concentrations for the first time. N-Chlorotaurine (12.5 to 50 μM) demonstrated a bactericidal effect i.e., a 2 to 4 log10 reduction in viable counts, after incubation at 37°C for 6 to 9 h at pH 7.0, which effect was significantly enhanced in an acidic milieu (at pH 5.0), with a 3 to 4 log10 reduction after 2 to 3 h. Moreover, bacteria were attenuated after being incubated in N-chlorotaurine for a sublethal time, as demonstrated with the mouse peritonitis model. The supernatant of stimulated granulocytes exhibited similar activity. Transmission electron microscopy revealed changes in the bacterial cell membrane and cytoplasmic disintegration with both reacting systems, even in the case of a mere attenuation. The results of this study suggest a significant bactericidal function of N-chlorotaurine and other chloramines during inflammation. PMID:10952603

Effects of ozone nano-bubble water on periodontopathic bacteria and oral cells - in vitro studies

NASA Astrophysics Data System (ADS)

Hayakumo, Sae; Arakawa, Shinichi; Takahashi, Masayoshi; Kondo, Keiko; Mano, Yoshihiro; Izumi, Yuichi

2014-10-01

The aims of the present study were to evaluate the bactericidal activity of a new antiseptic agent, ozone nano-bubble water (NBW3), against periodontopathogenic bacteria and to assess the cytotoxicity of NBW3 against human oral cells. The bactericidal activities of NBW3 against representative periodontopathogenic bacteria, Porphyromonas gingivalis (P. gingivalis) and Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans) were evaluated using in vitro time-kill assays. The cytotoxicity of NBW3 was evaluated using three-dimensional human buccal and gingival tissue models. The numbers of colony forming units (CFUs)/mL of P. gingivalis and A. actinomycetemcomitans exposed to NBW3 dropped to below the lower limit of detection (<10 CFUs mL-1) after only 0.5 min of exposure. There were only minor decreases in the viability of oral tissue cells after 24 h of exposure to NBW3. These results suggest that NBW3 possesses potent bactericidal activity against representative periodontopathogenic bacteria and is not cytotoxic to cells of human oral tissues. The use of NBW3 as an adjunct to periodontal therapy would be promising.

Antioxidant Effect of Melatonin on the Functional Activity of Colostral Phagocytes in Diabetic Women

PubMed Central

Fagundes, Danny L. G.; Calderon, Iracema M. P.; França, Eduardo L.

2013-01-01

Melatonin is involved in a number of physiological and oxidative processes, including functional regulation in human milk. The present study investigated the mechanisms of action of melatonin and its effects on the functional activity of colostral phagocytes in diabetic women. Colostrum samples were collected from normoglycemic (N = 38) and diabetic (N = 38) women. We determined melatonin concentration, superoxide release, bactericidal activity and intracellular Ca2+ release by colostral phagocytes treated or not with 8-(Diethylamino) octyl-3,4,5-trimethoxybenzoate hydrochloride (TMB-8) and incubated with melatonin and its precursor (N-acetyl-serotonin-NAS), antagonist (luzindole) and agonist (chloromelatonin-CMLT). Melatonin concentration was higher in colostrum samples from hyperglycemic than normoglycemic mothers. Melatonin stimulated superoxide release by colostral phagocytes from normoglycemic but not hyperglycemic women. NAS increased superoxide, irrespective of glycemic status, whereas CMTL increased superoxide only in cells from the normoglycemic group. Phagocytic activity in colostrum increased significantly in the presence of melatonin, NAS and CMLT, irrespective of glycemic status. The bactericidal activity of colostral phagocytes against enterophatogenic Escherichia coli (EPEC) increased in the presence of melatonin or NAS in the normoglycemic group, but not in the hyperglycemic group. Luzindole blocked melatonin action on colostrum phagocytes. Phagocytes from the normoglycemic group treated with melatonin exhibited an increase in intracellular Ca2+ release. Phagocytes treated with TMB-8 (intracellular Ca2+ inhibitor) decreased superoxide, bactericidal activity and intracellular Ca2+ release in both groups. The results obtained suggest an interactive effect of glucose metabolism and melatonin on colostral phagocytes. In colostral phagocytes from normoglycemic mothers, melatonin likely increases the ability of colostrum to protect against EPEC and other infections. In diabetic mothers, because maternal hyperglycemia modifies the functional activity of colostrum phagocytes, melatonin effects are likely limited to anti-inflammatory processes, with low superoxide release and bactericidal activity. PMID:23437270

Differential mechanisms of complement-mediated neutralization of the closely related paramyxoviruses simian virus 5 and mumps virus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Johnson, John B.; Capraro, Gerald A.; Parks, Griffith D.

2008-06-20

The complement system is an important component of the innate immune response to virus infection. The role of human complement pathways in the in vitro neutralization of three closely related paramyxoviruses, Simian Virus 5 (SV5), Mumps virus (MuV) and Human Parainfluenza virus type 2 (HPIV2) was investigated. Sera from ten donors showed high levels of neutralization against HPIV2 that was largely complement-independent, whereas nine of ten donor sera were found to neutralize SV5 and MuV only in the presence of active complement pathways. SV5 and MuV neutralization proceeded through the alternative pathway of the complement cascade. Electron microscopy studies andmore » biochemical analyses showed that treatment of purified SV5 with human serum resulted in C3 deposition on virions and the formation of massive aggregates, but there was relatively little evidence of virion lysis. Treatment of MuV with human serum also resulted in C3 deposition on virions, however in contrast to SV5, MuV particles were lysed by serum complement and there was relatively little aggregation. Assays using serum depleted of complement factors showed that SV5 and MuV neutralization in vitro was absolutely dependent on complement factor C3, but was not dependent on downstream complement factors C5 or C8. Our results indicate that even though antibodies exist that recognize both SV5 and MuV, they are mostly non-neutralizing and viral inactivation in vitro occurs through the alternative pathway of complement. The implications of our work for development of paramyxovirus vectors and vaccines are discussed.« less

Prevalence of factor H-binding protein variants and NadA among meningococcal group B isolates from the United States: implications for the development of a multicomponent group B vaccine.

PubMed

Beernink, Peter T; Welsch, Jo Anne; Harrison, Lee H; Leipus, Arunas; Kaplan, Sheldon L; Granoff, Dan M

2007-05-15

Two promising recombinant meningococcal protein vaccines are in development. One contains factor H-binding protein (fHBP) variants (v.) 1 and 2, whereas the other contains v.1 and 4 other antigens discovered by genome mining (5 component [5C]). Antibodies against fHBP are bactericidal against strains within a variant group. There are limited data on the prevalence of strains expressing different fHBP variants in the United States. A total of 143 group B isolates from patients hospitalized in the United States were tested for fHBP variant by quantitative polymerase chain reaction, for reactivity with 6 anti-fHBP monoclonal antibodies (MAb) by dot immunoblotting, and for susceptibility to bactericidal activity of mouse antisera. fHBP v.1 isolates predominated in California (83%), whereas isolates expressing v.1 (53%) or v.2 (42%) were common in 9 other states. Isolates representative of 5 anti-fHBP MAb-binding phenotypes (70% of isolates) were highly susceptible to anti-fHBP v.1 or v.2 bactericidal activity, whereas 3 phenotypes were approximately 50% susceptible. Collectively, antibodies against the fHBP v.1 and v.2 vaccine and the 5C vaccine killed 76% and 83% of isolates, respectively. Susceptibility to bactericidal activity can be predicted, in part, on the basis of fHBP phenotypes. Both vaccines have the potential to prevent most group B disease in the United States.

DNA Is an Antimicrobial Component of Neutrophil Extracellular Traps

PubMed Central

Halverson, Tyler W.R.; Wilton, Mike; Poon, Karen K. H.; Petri, Björn; Lewenza, Shawn

2015-01-01

Neutrophil extracellular traps (NETs) comprise an ejected lattice of chromatin enmeshed with granular and nuclear proteins that are capable of capturing and killing microbial invaders. Although widely employed to combat infection, the antimicrobial mechanism of NETs remains enigmatic. Efforts to elucidate the bactericidal component of NETs have focused on the role of NET-bound proteins including histones, calprotectin and cathepsin G protease; however, exogenous and microbial derived deoxyribonuclease (DNase) remains the most potent inhibitor of NET function. DNA possesses a rapid bactericidal activity due to its ability to sequester surface bound cations, disrupt membrane integrity and lyse bacterial cells. Here we demonstrate that direct contact and the phosphodiester backbone are required for the cation chelating, antimicrobial property of DNA. By treating NETs with excess cations or phosphatase enzyme, the antimicrobial activity of NETs is neutralized, but NET structure, including the localization and function of NET-bound proteins, is maintained. Using intravital microscopy, we visualized NET-like structures in the skin of a mouse during infection with Pseudomonas aeruginosa. Relative to other bacteria, P. aeruginosa is a weak inducer of NETosis and is more resistant to NETs. During NET exposure, we demonstrate that P. aeruginosa responds by inducing the expression of surface modifications to defend against DNA-induced membrane destabilization and NET-mediated killing. Further, we show induction of this bacterial response to NETs is largely due to the bacterial detection of DNA. Therefore, we conclude that the DNA backbone contributes both to the antibacterial nature of NETs and as a signal perceived by microbes to elicit host-resistance strategies. PMID:25590621

Staphylococcus epidermidis induces complement activation, tumor necrosis factor and interleukin-1, a shock-like state and tissue injury in rabbits without endotoxemia. Comparison to Escherichia coli.

PubMed Central

Wakabayashi, G; Gelfand, J A; Jung, W K; Connolly, R J; Burke, J F; Dinarello, C A

1991-01-01

Tumor necrosis factor (TNF) and IL-1 are thought to mediate many of the pathophysiologic changes of endotoxemia and Gram-negative bacteremia. In these studies, heat-killed Staphylococcus epidermidis were infused into rabbits to determine whether an endotoxin (LPS)-free microorganism also elicits cytokinemia and the physiologic abnormalities seen in Gram-negative bacteremia. S. epidermidis induced complement activation, circulating TNF and IL-1, and hypotension to the same degree as did one-twentieth the number of heat-killed Escherichia coli. Circulating IL-1 beta levels had a greater correlation coefficient (r = 0.81, P less than 0.001) with the degree of hypotension than TNF levels (r = 0.48, P less than 0.02). Leukopenia, thrombocytopenia, diffuse pulmonary capillary aggregation of neutrophils, and hepatic necrosis with neutrophil infiltration were observed to the same extent after either S. epidermidis or E. coli infusion. However, S. epidermidis infusion did not induce significant (less than 60 pg/ml) endotoxemia, whereas E. coli infusion resulted in high (11,000 pg/ml) serum endotoxin levels. S. epidermidis, E. coli, LPS, or S. epidermidis-derived lipoteichoic acid (LTA) induced TNF and IL-1 from blood mononuclear cells in vitro. E. coli organisms and LPS were at least 100-fold more potent than S. epidermidis or LTA. Thus, a shock-like state with similar levels of complement activation as well as circulating levels of IL-1 and TNF were observed following either S. epidermidis or E. coli. These data provide further evidence that host factors such as IL-1 and TNF are common mediators of the septic shock syndrome regardless of the organism. Images PMID:2040686

Ly6G-mediated depletion of neutrophils is dependent on macrophages.

PubMed

Bruhn, Kevin W; Dekitani, Ken; Nielsen, Travis B; Pantapalangkoor, Paul; Spellberg, Brad

2016-01-01

Antibody-mediated depletion of neutrophils is commonly used to study neutropenia. However, the mechanisms by which antibodies deplete neutrophils have not been well defined. We noticed that mice deficient in complement and macrophages had blunted neutrophil depletion in response to anti-Ly6G monoclonal antibody (MAb) treatment. In vitro, exposure of murine neutrophils to anti-Ly6G MAb in the presence of plasma did not result in significant depletion of cells, either in the presence or absence of complement. In vivo, anti-Ly6G-mediated neutrophil depletion was abrogated following macrophage depletion, but not complement depletion, indicating a requirement for macrophages to induce neutropenia by this method. These results inform the use and limitations of anti-Ly6G antibody as an experimental tool for depleting neutrophils in various immunological settings.

Homologous species restriction of the complement-mediated killing of nucleated cells.

PubMed Central

Yamamoto, H; Blaas, P; Nicholson-Weller, A; Hänsch, G M

1990-01-01

The homologous restriction of complement (C) lysis is attributed to membrane proteins: decay-accelerating factor (DAF), C8 binding protein (C8bp) and P18/CD59. Since these proteins are also expressed on peripheral blood cells, species restriction was tested for in the complement-mediated killing of antibody-coated human leucocytes by human or rabbit complement. Killing was more efficient when rabbit complement was used. Preincubation of cells with an antibody to DAF abolished the difference. When C1-7 sites were first attached to the cells and either rabbit or human C8, C9 were added, the killing of monocytes and lymphocytes was equally efficient; only in polymorphonuclear neutrophils was a higher efficiency of rabbit C8, C9 seen. Thus, in contrast to haemolysis, restriction occurred predominantly at the C3 level and the action of the terminal complement components was not inhibited. Since C8bp isolated from peripheral blood cells showed essentially similar characteristics as the erythrocyte-derived C8bp, the failure of C8bp to inhibit the action of the terminal components on nucleated cells might reflect differences of the complement membrane interactions between erythrocytes or nucleated cells, respectively. Images Figure 5 PMID:1697561

More than just immune evasion: Hijacking complement by Plasmodium falciparum.

PubMed

Schmidt, Christoph Q; Kennedy, Alexander T; Tham, Wai-Hong

2015-09-01

Malaria remains one of the world's deadliest diseases. Plasmodium falciparum is responsible for the most severe and lethal form of human malaria. P. falciparum's life cycle involves two obligate hosts: human and mosquito. From initial entry into these hosts, malaria parasites face the onslaught of the first line of host defence, the complement system. In this review, we discuss the complex interaction between complement and malaria infection in terms of hosts immune responses, parasite survival and pathogenesis of severe forms of malaria. We will focus on the role of complement receptor 1 and its associated polymorphisms in malaria immune complex clearance, as a mediator of parasite rosetting and as an entry receptor for P. falciparum invasion. Complement evasion strategies of P. falciparum parasites will also be highlighted. The sexual forms of the malaria parasites recruit the soluble human complement regulator Factor H to evade complement-mediated killing within the mosquito host. A novel evasion strategy is the deployment of parasite organelles to divert complement attack from infective blood stage parasites. Finally we outline the future challenge to understand the implications of these exploitation mechanisms in the interplay between successful infection of the host and pathogenesis observed in severe malaria. Copyright © 2015 Elsevier Ltd. All rights reserved.

Breaking down the complement system: a review and update on novel therapies.

PubMed

Reddy, Yuvaram N V; Siedlecki, Andrew M; Francis, Jean M

2017-03-01

The complement system represents one of the more primitive forms of innate immunity. It has increasingly been found to contribute to pathologies in the native and transplanted kidney. We provide a concise review of the physiology of the complement cascade, and discuss current and upcoming complement-based therapies. Current agents in clinical use either bind to complement components directly or prevent complement from binding to antibodies affixed to the endothelial surface. These include C1 esterase inhibitors, anti-C5 mAbs, anti-CD20 mAbs, and proteasome inhibitors. Treatment continues to show efficacy in the atypical hemolytic uremic syndrome and antibody-mediated rejection. Promising agents not currently available include CCX168, TP10, AMY-101, factor D inhibitors, coversin, and compstatin. Several new trials are targeting complement inhibition to treat antineutrophilic cystoplasmic antibody (ANCA)-associated vasculitis, C3 glomerulopathy, thrombotic microangiopathy, and IgA nephropathy. New agents for the treatment of the atypical hemolytic uremic syndrome are also in development. Complement-based therapies are being considered for targeted therapy in the atypical hemolytic uremic syndrome and antibody-mediated rejection, C3 glomerulopathy, and ANCA-associated vasculitis. A few agents are currently in use as orphan drugs. A number of other drugs are in clinical trials and, overall, are showing promising preliminary results.

[Age changes of immunological, morphological and biochemical indices of male reproductive system].

PubMed

Boĭko, O V; Akhmineeva, A Kh; Gudinskaia, N I; Boĭko, V I; Kozak, D M

2014-01-01

The article analyzes the dependence of bactericidal activity of sperm--natural resistance factors controlling the survival of bacteria in the urogenital tract, on the age of men. These data are compared with the results of the standard (on the recommendations of the WHO) spermogram, reflecting reproductive health. Due to the fact that one of the main etiological agents of infectious disease groups in the male reproductive system in adulthood are Staphylococcus spp., we consider the level of bactericidal activity of sperm in resident and transient carriage of S. aureus and S. epidermidis.

Microstructural, spectroscopic, and antibacterial properties of silver-based hybrid nanostructures biosynthesized using extracts of coriander leaves and seeds

PubMed Central

Luna, Carlos; Barriga-Castro, Enrique Díaz; Gómez-Treviño, Alberto; Núñez, Nuria O; Mendoza-Reséndez, Raquel

2016-01-01

Coriander leaves and seeds have been highly appreciated since ancient times, not only due to their pleasant flavors but also due to their inhibitory activity on food degradation and their beneficial properties for health, both ascribed to their strong antioxidant activity. Recently, it has been shown that coriander leaf extracts can mediate the synthesis of metallic nanoparticles through oxidation/reduction reactions. In the present study, extracts of coriander leaves and seeds have been used as reaction media for the wet chemical synthesis of ultrafine silver nanoparticles and nanoparticle clusters, with urchin- and tree-like shapes, coated by biomolecules (mainly, proteins and polyphenols). In this greener route of nanostructure preparation, the active biocompounds of coriander simultaneously play the roles of reducing and stabilizing agents. The morphological and microstructural studies of the resulting biosynthesized silver nanostructures revealed that the nanostructures prepared with a small concentration of the precursor Ag salt (AgNO3 =5 mM) exhibit an ultrafine size and a narrow size distribution, whereas particles synthesized with high concentrations of the precursor Ag salt (AgNO3 =0.5 M) are polydisperse and formation of supramolecular structures occurs. Fourier transform infrared and Raman spectroscopy studies indicated that the bioreduction of the Ag− ions takes place through their interactions with free amines, carboxylate ions, and hydroxyl groups. As a consequence of such interactions, residues of proteins and polyphenols cap the biosynthesized Ag nanoparticles providing them a hybrid core/shell structure. In addition, these biosynthesized Ag nanomaterials exhibited size-dependent plasmon extinction bands and enhanced bactericidal activities against both Gram-positive and Gram-negative bacteria, displaying minimal inhibitory Ag concentrations lower than typical values reported in the literature for Ag nanoparticles, probably due to the synergy of the bactericidal activities of the Ag nanoparticle cores and their capping ligands. PMID:27703347

influence of TEM-1 beta-lactamase on the pharmacodynamic activity of simulated total versus free-drug serum concentrations of cefditoren (400 milligrams) versus amoxicillin-clavulanic acid (2,000/125 milligrams) against Haemophilus influenzae strains exhibiting an N526K mutation in the ftsI gene.

PubMed

Torrico, M; Aguilar, L; González, N; Giménez, M J; Echeverría, O; Cafini, F; Sevillano, D; Alou, L; Coronel, P; Prieto, J

2007-10-01

The aim of this study was to explore bactericidal activity of total and free serum simulated concentrations after the oral administration of cefditoren (400 mg, twice daily [bid]) versus the oral administration of amoxicillin-clavulanic acid extended release formulation (2,000/125 mg bid) against Haemophilus influenzae. A computerized pharmacodynamic simulation was performed, and colony counts and beta-lactamase activity were determined over 48 h. Three strains were used: ampicillin-susceptible, beta-lactamase-negative ampicillin-resistant (BLNAR) (also resistant to amoxicillin-clavulanic acid) and beta-lactamase-positive amoxicillin-clavulanic acid-resistant (BLPACR) strains, with cefditoren MICs of < or =0.12 microg/ml and amoxicillin-clavulanic acid MICs of 2, 8, and 8 microg/ml, respectively. Against the ampicillin-susceptible and BLNAR strains, bactericidal activity (> or =3 log(10) reduction) was obtained from 6 h on with either total and free cefditoren or amoxicillin-clavulanic acid. Against the BLPACR strain, free cefditoren showed bactericidal activity from 8 h on. In amoxicillin-clavulanic acid simulations the increase in colony counts from 4 h on occurred in parallel with the increase in beta-lactamase activity for the BLPACR strain. Since both BLNAR and BLPACR strains exhibited the same MIC, this was due to the significantly lower (P < or = 0.012) amoxicillin concentrations from 4 h on in simulations with beta-lactamase positive versus negative strains, thus decreasing the time above MIC (T>MIC). From a pharmacodynamic point of view, the theoretical amoxicillin T>MIC against strains with elevated ampicillin/amoxicillin-clavulanic acid MICs should be considered with caution since the presence of beta-lactamase inactivates the antibiotic, thus rendering inaccurate theoretical calculations. The experimental bactericidal activity of cefditoren is maintained over the dosing interval regardless of the presence of a mutation in the ftsI gene or beta-lactamase production.

The bactericidal activity of glutaraldehyde-impregnated polyurethane.

PubMed

Sehmi, Sandeep K; Allan, Elaine; MacRobert, Alexander J; Parkin, Ivan

2016-10-01

Although glutaraldehyde is known to be bactericidal in solution, its potential use to create novel antibacterial polymers suitable for use in healthcare environments has not been evaluated. Here, novel materials were prepared in which glutaraldehyde was either incorporated into polyurethane using a simple "swell-encapsulation-shrink" method (hereafter referred to as "glutaraldehyde-impregnated polyurethane"), or simply applied to the polymer surface (hereafter referred to as "glutaraldehyde-coated polyurethane"). The antibacterial activity of glutaraldehyde-impregnated and glutaraldehyde-coated polyurethane samples was tested against Escherichia coli and Staphylococcus aureus. Glutaraldehyde-impregnated polyurethane resulted in a 99.9% reduction in the numbers of E. coli within 2 h and a similar reduction of S. aureus within 1 h, whereas only a minimal reduction in bacterial numbers was observed when the biocide was bound to the polymer surface. After 15 days, however, the bactericidal activity of the impregnated material was substantially reduced presumably due to polymerization of glutaraldehyde. Thus, although glutaraldehyde retains antibacterial activity when impregnated into polyurethane, activity is not maintained for extended periods of time. Future work should examine the potential of chemical modification of glutaraldehyde and/or polyurethane to improve the useful lifespan of this novel antibacterial polymer. © 2016 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Superhydrophilic nanopillar-structured quartz surfaces for the prevention of biofilm formation in optical devices

NASA Astrophysics Data System (ADS)

Han, Soo; Ji, Seungmuk; Abdullah, Abdullah; Kim, Duckil; Lim, Hyuneui; Lee, Donghyun

2018-01-01

Bacterial biofilm formation on optical devices such as contact lenses, optical glasses, endoscopic devices, and microscopic slides and lenses are major concerns in the field of medicine and biomedical engineering. To solve these problems, here we present the first report of superhydrophilic transparent nanopillar-structured surfaces with bactericidal properties. To construct bactericidal surfaces, we imitated a topological mechanism found in nature in which nanopillar-structured surfaces cause a mechanical disruption of the outer cell membranes of bacteria, resulting in bacterial cell death. We used nanosphere lithography to fabricate nanopillars with various sharpnesses and heights on a quartz substrate. Water contact angle and light reflectance measurements revealed superhydrophilic, antifogging and antireflective properties, which are important for use in optical devices. To determine bactericidal efficiency, the fabricated surfaces were incubated and tested against two Gram-negative bacteria associated with biofilm formation and various diseases in humans, Pseudomonas aeruginosa and Escherichia coli. The highest bactericidal activity was achieved with nanopillars that measured 300 nm in height and 10 nm in apex diameter. Quartz substrates patterned with such nanopillars killed ∼38,000 P. aeruginosa and ∼27,000 E. coli cells cm-2 min-1, respectively. Thus, the newly designed nanopillar-structured bactericidal surfaces are suitable for use in the development of superhydrophilic and transparent optical devices.

Complement anaphylatoxins as immune regulators in cancer.

PubMed

Sayegh, Eli T; Bloch, Orin; Parsa, Andrew T

2014-08-01

The role of the complement system in innate immunity is well characterized. However, a recent body of research implicates the complement anaphylatoxins C3a and C5a as insidious propagators of tumor growth and progression. It is now recognized that certain tumors elaborate C3a and C5a and that complement, as a mediator of chronic inflammation and regulator of immune function, may in fact foster rather than defend against tumor growth. A putative mechanism for this function is complement-mediated suppression of immune effector cells responsible for immunosurveillance within the tumor microenvironment. This paradigm accords with models of immune dysregulation, such as autoimmunity and infectious disease, which have defined a pathophysiological role for abnormal complement signaling. Several types of immune cells express the cognate receptors for the complement anaphylatoxins, C3aR and C5aR, and demonstrate functional modulation in response to complement stimulation. In turn, impairment of antitumor immunity has been intimately tied to tumor progression in animal models of cancer. In this article, the literature was systematically reviewed to identify studies that have characterized the effects of the complement anaphylatoxins on the composition and function of immune cells within the tumor microenvironment. The search identified six studies based upon models of lymphoma and ovarian, cervical, lung, breast, and mammary cancer, which collectively support the paradigm of complement as an immune regulator in the tumor microenvironment. © 2014 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

Antibacterial applications of α-Fe2O3/Co3O4 nanocomposites and study of their structural, optical, magnetic and cytotoxic characteristics

NASA Astrophysics Data System (ADS)

Bhushan, Mayank; Kumar, Yogesh; Periyasamy, Latha; Viswanath, Annamraju Kasi

2018-02-01

Owing to their multiple mechanisms of bactericidal activity, inorganic metal oxides and hybrid metal oxide nanocomposites may serve as a new class of effective disinfectants. Among metal oxide nanoparticles, iron oxide nanoparticles exhibit minimal or no cytotoxicity to human cells with very efficient bactericidal properties over a wide spectrum of bacteria. This paper presents the very first report on antibacterial properties of novel nanocomposites of iron oxide and cobalt oxide nanoparticles against pathogenic bacterial strains B. subtilis, S. aureus, E.coli and S. typhi. The enhanced bactericidal activity of the Fe/Co oxide nanocomposite was the result of synergistic effect of iron oxide and cobalt oxide nanoparticles. The nanocomposites were synthesized using co-precipitation route with increasing cobalt content in the sample and further characterized using XRD, TEM, Raman and VSM to investigate structural, optical and magnetic properties of the prepared nanocomposites, respectively. Also, the prepared nanocomposites were highly biocompatible and found non-toxic to human cell line MCF7.

In vitro growth inhibition of mastitis causing bacteria by phenolics and metal chelators

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chew, B.P.; Tjoelker, L.W.; Tanaka, T.S.

1985-11-01

Antimicrobial activities of three phenolic compounds and four metal chelators were tested at 0, 250, 500, and 1000 ppm in vitro against four major mastitis-causing bacteria, Streptococcus agalactiae, Staphylococcus aureus, Klebsiella pnuemoniae, and Escherichia coli. Overall, butylated hydroxyanisole and tert-butylhydroquinone showed the greatest antimicrobial activity. These phenolics were bactericidal at 250 to 500 ppm against all four bacteria tested. The butylated hydroxytoluene was bactericidal against the gram-positive bacteria but was ineffective against the coliforms. At 250 ppm, disodium ethylenediaminetetraacetic acid was bactericidal against the gram-positive bacteria but much less effective against the gram-negatives. However, diethylene-triaminepentaacetic acid was more growth inhibitorymore » than ethylenediaminetetraacetic acid against the gram-negative bacteria and especially against Escherichia coli. All other compounds were generally much less effective or ineffective against all four microorganisms. Therefore, butylated hydroxyanisole, butylated hydroxytoluene, tert-butylhydroquinone, ethylenediaminetetraacetic acid, and diethylenetriaminepentaacetic acid may have practical implications in the prevention or treatment of bovine mastitis.« less

Serum bactericidal activities of moxifloxacin and levofloxacin against aerobic and anaerobic intra-abdominal pathogens.

PubMed

Stein, Gary E; Schooley, Sharon; Tyrrell, Kerin L; Citron, Diane M; Nicolau, David P; Goldstein, Ellie J C

2008-02-01

We studied the serum bactericidal activity (SBA) of moxifloxacin and levofloxacin against common pathogens associated with complicated intra-abdominal infections. Ten healthy volunteers received a single dose of moxifloxacin (400 mg) and levofloxacin (750 mg) and serum samples were collected at 2, 4, 8, 12, and 24h after the dose of each drug. Bactericidal titers in serum over time were determined for aerobic gram-negative bacilli (Escherichia coli, Klebseilla pneumoniae, and Enterobacter cloacae) and anaerobic bacteria (Bacteroides fragilis, Bacteroides thetaiotaomicron, Prevotella bivia, and Finegoldia magna). Both fluoroquinolones provided rapid (2h) attainment and prolonged (24h) SBA (titers > or = 1:8) against each of the aerobic bacilli studied. SBA was observed for at least 12h against B. fragilis strains with MICs < or = 2 microg/ml to moxifloxacin and < or = 4 microg/ml to levofloxacin. Prolonged (12h) SBA (titers > or = 1:2) was also observed against isolates of B. thetaiotaomicron, P. bivia, and F. magna with moxifloxacin < or = MICs 2 microg/ml.

Pathogen- and host-directed anti-inflammatory activities of macrolide antibiotics.

PubMed

Steel, Helen C; Theron, Annette J; Cockeran, Riana; Anderson, Ronald; Feldman, Charles

2012-01-01

Macrolide antibiotics possess several, beneficial, secondary properties which complement their primary antimicrobial activity. In addition to high levels of tissue penetration, which may counteract seemingly macrolide-resistant bacterial pathogens, these agents also possess anti-inflammatory properties, unrelated to their primary antimicrobial activity. Macrolides target cells of both the innate and adaptive immune systems, as well as structural cells, and are beneficial in controlling harmful inflammatory responses during acute and chronic bacterial infection. These secondary anti-inflammatory activities of macrolides appear to be particularly effective in attenuating neutrophil-mediated inflammation. This, in turn, may contribute to the usefulness of these agents in the treatment of acute and chronic inflammatory disorders of both microbial and nonmicrobial origin, predominantly of the airways. This paper is focused on the various mechanisms of macrolide-mediated anti-inflammatory activity which target both microbial pathogens and the cells of the innate and adaptive immune systems, with emphasis on their clinical relevance.

Pathogen- and Host-Directed Anti-Inflammatory Activities of Macrolide Antibiotics

PubMed Central

Steel, Helen C.; Theron, Annette J.; Cockeran, Riana; Anderson, Ronald; Feldman, Charles

2012-01-01

Macrolide antibiotics possess several, beneficial, secondary properties which complement their primary antimicrobial activity. In addition to high levels of tissue penetration, which may counteract seemingly macrolide-resistant bacterial pathogens, these agents also possess anti-inflammatory properties, unrelated to their primary antimicrobial activity. Macrolides target cells of both the innate and adaptive immune systems, as well as structural cells, and are beneficial in controlling harmful inflammatory responses during acute and chronic bacterial infection. These secondary anti-inflammatory activities of macrolides appear to be particularly effective in attenuating neutrophil-mediated inflammation. This, in turn, may contribute to the usefulness of these agents in the treatment of acute and chronic inflammatory disorders of both microbial and nonmicrobial origin, predominantly of the airways. This paper is focused on the various mechanisms of macrolide-mediated anti-inflammatory activity which target both microbial pathogens and the cells of the innate and adaptive immune systems, with emphasis on their clinical relevance. PMID:22778497

Minimization of bacterial size allows for complement evasion and is overcome by the agglutinating effect of antibody

PubMed Central

Dalia, Ankur B.; Weiser, Jeffrey N.

2011-01-01

SUMMARY The complement system, which functions by lysing pathogens directly or by promoting their uptake by phagocytes, is critical for controlling many microbial infections. Here we show that in Streptococcus pneumoniae, increasing bacterial chain length sensitizes this pathogen to complement deposition and subsequent uptake by human neutrophils. Consistent with this, we show that minimizing chain length provides wild-type bacteria with a competitive advantage in vivo in a model of systemic infection. Investigating how the host overcomes this virulence strategy, we find that antibody promotes complement-dependent opsonophagocytic killing of Streptococcus pneumoniae and lysis of Haemophilus influenzae independent of Fc-mediated effector functions. Consistent with the agglutinating effect of antibody, F(ab′)2 but not Fab could promote this effect. Therefore, increasing pathogen size, whether by natural changes in cellular morphology or via antibody-mediated agglutination, promotes complement-dependent killing. These observations have broad implications for how cell size and morphology can affect virulence among pathogenic microbes. PMID:22100164

Shielding of a lipooligosaccharide IgM epitope allows evasion of neutrophil-mediated killing of an invasive strain of nontypeable Haemophilus influenzae.

PubMed

Langereis, Jeroen D; Weiser, Jeffrey N

2014-07-22

Nontypeable Haemophilus influenzae is a frequent cause of noninvasive mucosal inflammatory diseases but may also cause invasive diseases, such as sepsis and meningitis, especially in children and the elderly. Infection by nontypeable Haemophilus influenzae is characterized by recruitment of neutrophilic granulocytes. Despite the presence of a large number of neutrophils, infections with nontypeable Haemophilus influenzae are often not cleared effectively by the antimicrobial activity of these immune cells. Herein, we examined how nontypeable Haemophilus influenzae evades neutrophil-mediated killing. Transposon sequencing (Tn-seq) was used on an isolate resistant to neutrophil-mediated killing to identify genes required for its survival in the presence of human neutrophils and serum, which provided a source of complement and antibodies. Results show that nontypeable Haemophilus influenzae prevents complement-dependent neutrophil-mediated killing by expression of surface galactose-containing oligosaccharide structures. These outer-core structures block recognition of an inner-core lipooligosaccharide epitope containing glucose attached to heptose HepIII-β1,2-Glc by replacement with galactose attached to HepIII or through shielding HepIII-β1,2-Glc by phase-variable attachment of oligosaccharide chain extensions. When the HepIII-β1,2-Glc-containing epitope is expressed and exposed, nontypeable Haemophilus influenzae is opsonized by naturally acquired IgM generally present in human serum and subsequently phagocytosed and killed by human neutrophils. Clinical nontypeable Haemophilus influenzae isolates containing galactose attached to HepIII that are not recognized by this IgM are more often found to cause invasive infections. Importance: Neutrophils are white blood cells that specialize in killing pathogens and are recruited to sites of inflammation. However, despite the presence of large numbers of neutrophils in the middle ear cavity and lungs of patients with otitis media or chronic obstructive pulmonary disease, respectively, the bacterium nontypeable Haemophilus influenzae is often not effectively cleared from these locations by these immune cells. In order to understand how nontypeable Haemophilus influenzae is able to cause inflammatory diseases in the presence of neutrophils, we determined the mechanism that underlies resistance to neutrophil-mediated killing. We have shown that nontypeable Haemophilus influenzae prevents binding of antibodies of the IgM subtype through changes in their surface lipooligosaccharide structure, thereby preventing complement activation and clearance by human neutrophils. Copyright © 2014 Langereis and Weiser.

Inactivation of Candida glabrata by a humid DC argon discharge afterglow: dominant contributions of short-lived aqueous active species

NASA Astrophysics Data System (ADS)

Xiong, Qing; Liu, Hongbin; Lu, Weiping; Chen, Qiang; Xu, Le; Wang, Xia; Zhu, Qunlin; Zeng, Xue; Yi, Ping

2017-05-01

Plasma medicine applications are currently attracting significant interest all over the world. Bactericidal treatments of Candida glabrata cultured in saline suspension are performed in this study by a room-temperature reactive afterglow of a DC-driven argon discharge. Water vapor was added to the discharge to study the inactivation contributions of reactive hydrolytic species including OH and H2O2 transporting along the gas flow to the treated solutions. The inactivation results indicate that the dominant roles in the bactericidal treatments are played by the short-lived aqueous active species, but not the stable species like H2O2aq (aq indicates an aqueous species). Further analysis shows that the ·OHaq radicals play an important role in the inactivation process. The ·OHaq radicals in the suspension are mostly produced from the direct dissolution of the OH species in the reactive afterglow. With the increase of added water vapor content, the ·OHaq production increases and enhances the inactivation efficiency of C. glabrata. Furthermore, it is found that the ambient air diffusion shows essential effects on the bactericidal activity of the remote humid argon discharge. Higher bactericidal effects can be obtained in open-space treatments compared to in a controlled Ar + H2O gas atmosphere. Key active air-byproduct species are believed to be generated in the suspension during the treatments and contributing to the inactivation process. Based on chemical analysis, the peroxynitrous acid ONOOHaq is considered as the key antimicrobial air-byproduct species. These results indicate the important dependence of plasma biomedical effects on the processing environment, which finally relates to the critical contributions of the key reactive species formed therein.

Membrane-Active Epithelial Keratin 6A Fragments (KAMPs) Are Unique Human Antimicrobial Peptides with a Non-αβ Structure

PubMed Central

Lee, Judy T. Y.; Wang, Guangshun; Tam, Yu Tong; Tam, Connie

2016-01-01

Antibiotic resistance is a pressing global health problem that threatens millions of lives each year. Natural antimicrobial peptides and their synthetic derivatives, including peptoids and peptidomimetics, are promising candidates as novel antibiotics. Recently, the C-terminal glycine-rich fragments of human epithelial keratin 6A were found to have bactericidal and cytoprotective activities. Here, we used an improved 2-dimensional NMR method coupled with a new protocol for structural refinement by low temperature simulated annealing to characterize the solution structure of these kerain-derived antimicrobial peptides (KAMPs). Two specific KAMPs in complex with membrane mimicking sodium dodecyl sulfate (SDS) micelles displayed amphipathic conformations with only local bends and turns, and a central 10-residue glycine-rich hydrophobic strip that is central to bactericidal activity. To our knowledge, this is the first report of non-αβ structure for human antimicrobial peptides. Direct observation of Staphylococcus aureus and Pseudomonas aeruginosa by scanning and transmission electron microscopy showed that KAMPs deformed bacterial cell envelopes and induced pore formation. Notably, in competitive binding experiments, KAMPs demonstrated binding affinities to LPS and LTA that did not correlate with their bactericidal activities, suggesting peptide-LPS and peptide-LTA interactions are less important in their mechanisms of action. Moreover, immunoprecipitation of KAMPs-bacterial factor complexes indicated that membrane surface lipoprotein SlyB and intracellular machineries NQR sodium pump and ribosomes are potential molecular targets for the peptides. Results of this study improve our understanding of the bactericidal function of epithelial cytokeratin fragments, and highlight an unexplored class of human antimicrobial peptides, which may serve as non-αβ peptide scaffolds for the design of novel peptide-based antibiotics. PMID:27891122

Treatment of platelets with riboflavin and ultraviolet light mediates complement activation and suppresses monocyte interleukin-12 production in whole blood.

PubMed

Loh, Y S; Dean, M M; Johnson, L; Marks, D C

2015-11-01

Pathogen inactivation (PI) and storage may alter the immunomodulatory capacity of platelets (PLTs). The aim of this study was to examine the effect of PI (Riboflavin and ultraviolet light treatment) and storage on the capacity of PLTs to induce cytokine responses in recipient inflammatory cells. A pool and split design was used to prepare untreated and PI-treated buffy coat-derived platelet concentrates (PCs). Samples were taken on days 2 and 7 postcollection and incubated with ABO/RhD-matched fresh whole blood for 6 h with or without lipopolysaccharide (LPS). The intracellular production of IP-10, MCP-1, MIP-1α, IL-8, IL-6, IL-10, IL-12, TNF-α and MIP-1β in monocytes and neutrophils was assessed using flow cytometry. Complement proteins in PLT supernatants were measured using a cytometric bead array. PLTs and PLT supernatant (both untreated and PI-treated) resulted in modulation of intracellular MIP-1β and IL-12 production in monocytes. Compared to untreated PLTs, PI-treated PLTs resulted in significantly lower LPS-induced monocyte IL-12 production (day 7). The concentration of C3a and C5a (and their desArg forms) was significantly increased in PLT supernatants following PI. PI results in decreased LPS-induced monocyte IL-12 production and increased complement activation. The association between platelet-induced complement activation and IL-12 production warrants further investigation. © 2015 International Society of Blood Transfusion.

Immediate memory consequences of the effect of emotion on attention to pictures.

PubMed

Talmi, Deborah; Anderson, Adam K; Riggs, Lily; Caplan, Jeremy B; Moscovitch, Morris

2008-03-01

Emotionally arousing stimuli are at once both highly attention grabbing and memorable. We examined whether emotional enhancement of memory (EEM) reflects an indirect effect of emotion on memory, mediated by enhanced attention to emotional items during encoding. We tested a critical prediction of the mediation hypothesis-that regions conjointly activated by emotion and attention would correlate with subsequent EEM. Participants were scanned with fMRI while they watched emotional or neutral pictures under instructions to attend to them a lot or a little, and were then given an immediate recognition test. A region in the left fusiform gyrus was activated by emotion, voluntary attention, and subsequent EEM. A functional network, different for each attention condition, connected this region and the amygdala, which was associated with emotion and EEM, but not with voluntary attention. These findings support an indirect cortical mediation account of immediate EEM that may complement a direct modulation model.

Immediate memory consequences of the effect of emotion on attention to pictures

PubMed Central

Talmi, Deborah; Anderson, Adam K.; Riggs, Lily; Caplan, Jeremy B.; Moscovitch, Morris

2008-01-01

Emotionally arousing stimuli are at once both highly attention grabbing and memorable. We examined whether emotional enhancement of memory (EEM) reflects an indirect effect of emotion on memory, mediated by enhanced attention to emotional items during encoding. We tested a critical prediction of the mediation hypothesis—that regions conjointly activated by emotion and attention would correlate with subsequent EEM. Participants were scanned with fMRI while they watched emotional or neutral pictures under instructions to attend to them a lot or a little, and were then given an immediate recognition test. A region in the left fusiform gyrus was activated by emotion, voluntary attention, and subsequent EEM. A functional network, different for each attention condition, connected this region and the amygdala, which was associated with emotion and EEM, but not with voluntary attention. These findings support an indirect cortical mediation account of immediate EEM that may complement a direct modulation model. PMID:18323572

Complement C5a-C5aR interaction enhances MAPK signaling pathway activities to mediate renal injury in trichloroethylene sensitized BALB/c mice.

PubMed

Zhang, Jia-xiang; Zha, Wan-sheng; Ye, Liang-ping; Wang, Feng; Wang, Hui; Shen, Tong; Wu, Chang-hao; Zhu, Qi-xing

2016-02-01

We have previously shown complement activation as a possible mechanism for trichloroethylene (TCE) sensitization, leading to multi-organ damage including the kidneys. In particular, excessive deposition of C5 and C5b-9-the membrane attack complex, which can generate significant tissue damage, was observed in the kidney tissue after TCE sensitization. The present study tested the hypothesis that anaphylatoxin C5a binding to its receptor C5aR mediates renal injury in TCE-sensitized BALB/c mice. BALB/c mice were sensitized through skin challenge with TCE, with or without pretreatment by the C5aR antagonist W54011. Kidney histopathology and the renal functional test were performed to assess renal injury, and immunohistochemistry and fluorescent labeling were carried out to assess C5a and C5aR expressions. TCE sensitization up-regulated C5a and C5aR expressions in kidney tissue, generated inflammatory infiltration, renal tubule damage, glomerular hypercellularity and impaired renal function. Antagonist pretreatment blocked C5a binding to C5aR and attenuated TCE-induced tissue damage and renal dysfunction. TCE sensitization also caused the deposition of major pro-inflammatory cytokines IL-2, TNF-α and IFN-γ in the kidney tissue (P < 0.05); this was accompanied by increased expression of P-p38, P-ERK and P-JNK proteins (P < 0.05). Pretreatment with the C5aR antagonist attenuated the increase of expression of P-p38, P-ERK and P-JNK proteins (P < 0.05) and also consistently reduced the TCE sensitization-induced increase of IL-2, TNF-α and IFN-γ (P < 0.05). These data identify C5a binding to C5aR, MAP kinase activation, and inflammatory cytokine release as a novel mechanism for complement-mediated renal injury by sensitization with TCE or other environmental chemicals. Copyright © 2015 John Wiley & Sons, Ltd.

Antimicrobial Testing Methods & Procedures: MB-26-02

EPA Pesticide Factsheets

Information about ATMP - SOP Neutralization of Microbicidal Activity using the OECD Quantitative Method for Evaluating Bactericidal Activity of Microbicides Used on Hard, Non-Porous Surfaces - MB-26-02

Antipneumococcal activity of ceftobiprole, a novel broad-spectrum cephalosporin.

PubMed

Kosowska, Klaudia; Hoellman, Dianne B; Lin, Gengrong; Clark, Catherine; Credito, Kim; McGhee, Pamela; Dewasse, Bonifacio; Bozdogan, Bülent; Shapiro, Stuart; Appelbaum, Peter C

2005-05-01

Ceftobiprole (previously known as BAL9141), an anti-methicillin-resistant Staphylococcus aureus cephalosporin, was very highly active against a panel of 299 drug-susceptible and -resistant pneumococci, with MIC(50) and MIC(90) values (microg/ml) of 0.016 and 0.016 (penicillin susceptible), 0.06 and 0.5 (penicillin intermediate), and 0.5 and 1.0 (penicillin resistant). Ceftobiprole, imipenem, and ertapenem had lower MICs against all pneumococcal strains than amoxicillin, cefepime, ceftriaxone, cefotaxime, cefuroxime, or cefdinir. Macrolide and penicillin G MICs generally varied in parallel, whereas fluoroquinolone MICs did not correlate with penicillin or macrolide susceptibility or resistance. All strains were susceptible to linezolid, quinupristin-dalfopristin, daptomycin, vancomycin, and teicoplanin. Time-kill analyses showed that at 1x and 2x the MIC, ceftobiprole was bactericidal against 10/12 and 11/12 strains, respectively. Levofloxacin, moxifloxacin, vancomycin, and teicoplanin were each bactericidal against 10 to 12 strains at 2x the MIC. Azithromycin and clarithromycin were slowly bactericidal, and telithromycin was bactericidal against only 5/12 strains at 2x the MIC. Linezolid was mainly bacteriostatic, whereas quinupristin-dalfopristin and daptomycin showed marked killing at early time periods. Prolonged serial passage in the presence of subinhibitory concentrations of ceftobiprole failed to yield mutants with high MICs towards this cephalosporin, and single-passage selection showed very low frequencies of spontaneous mutants with breakthrough MICs towards ceftobiprole.

Antipneumococcal Activity of Ceftobiprole, a Novel Broad-Spectrum Cephalosporin

PubMed Central

Kosowska, Klaudia; Hoellman, Dianne B.; Lin, Gengrong; Clark, Catherine; Credito, Kim; McGhee, Pamela; Dewasse, Bonifacio; Bozdogan, Bülent; Shapiro, Stuart; Appelbaum, Peter C.

2005-01-01

Ceftobiprole (previously known as BAL9141), an anti-methicillin-resistant Staphylococcus aureus cephalosporin, was very highly active against a panel of 299 drug-susceptible and -resistant pneumococci, with MIC50 and MIC90 values (μg/ml) of 0.016 and 0.016 (penicillin susceptible), 0.06 and 0.5 (penicillin intermediate), and 0.5 and 1.0 (penicillin resistant). Ceftobiprole, imipenem, and ertapenem had lower MICs against all pneumococcal strains than amoxicillin, cefepime, ceftriaxone, cefotaxime, cefuroxime, or cefdinir. Macrolide and penicillin G MICs generally varied in parallel, whereas fluoroquinolone MICs did not correlate with penicillin or macrolide susceptibility or resistance. All strains were susceptible to linezolid, quinupristin-dalfopristin, daptomycin, vancomycin, and teicoplanin. Time-kill analyses showed that at 1× and 2× the MIC, ceftobiprole was bactericidal against 10/12 and 11/12 strains, respectively. Levofloxacin, moxifloxacin, vancomycin, and teicoplanin were each bactericidal against 10 to 12 strains at 2× the MIC. Azithromycin and clarithromycin were slowly bactericidal, and telithromycin was bactericidal against only 5/12 strains at 2× the MIC. Linezolid was mainly bacteriostatic, whereas quinupristin-dalfopristin and daptomycin showed marked killing at early time periods. Prolonged serial passage in the presence of subinhibitory concentrations of ceftobiprole failed to yield mutants with high MICs towards this cephalosporin, and single-passage selection showed very low frequencies of spontaneous mutants with breakthrough MICs towards ceftobiprole. PMID:15855516

Selective bactericidal activity of nanopatterned superhydrophobic cicada Psaltoda claripennis wing surfaces.

PubMed

Hasan, Jafar; Webb, Hayden K; Truong, Vi Khanh; Pogodin, Sergey; Baulin, Vladimir A; Watson, Gregory S; Watson, Jolanta A; Crawford, Russell J; Ivanova, Elena P

2013-10-01

The nanopattern on the surface of Clanger cicada (Psaltoda claripennis) wings represents the first example of a new class of biomaterials that can kill bacteria on contact based solely on its physical surface structure. As such, they provide a model for the development of novel functional surfaces that possess an increased resistance to bacterial contamination and infection. Their effectiveness against a wide spectrum of bacteria, however, is yet to be established. Here, the bactericidal properties of the wings were tested against several bacterial species, possessing a range of combinations of morphology and cell wall type. The tested species were primarily pathogens, and included Bacillus subtilis, Branhamella catarrhalis, Escherichia coli, Planococcus maritimus, Pseudomonas aeruginosa, Pseudomonas fluorescens, and Staphylococcus aureus. The wings were found to consistently kill Gram-negative cells (i.e., B. catarrhalis, E. coli, P. aeruginosa, and P. fluorescens), while Gram-positive cells (B. subtilis, P. maritimus, and S. aureus) remained resistant. The morphology of the cells did not appear to play any role in determining cell susceptibility. The bactericidal activity of the wing was also found to be quite efficient; 6.1 ± 1.5 × 10(6) P. aeruginosa cells in suspension were inactivated per square centimeter of wing surface after 30-min incubation. These findings demonstrate the potential for the development of selective bactericidal surfaces incorporating cicada wing nanopatterns into the design.

The membrane attack complex of complement contributes to plasmin-induced synthesis of platelet-activating factor by endothelial cells and neutrophils

PubMed Central

Lupia, Enrico; Del Sorbo, Lorenzo; Bergerone, Serena; Emanuelli, Giorgio; Camussi, Giovanni; Montrucchio, Giuseppe

2003-01-01

Thrombolytic agents, used to restore blood flow to ischaemic tissues, activate several enzymatic systems with pro-inflammatory effects, thus potentially contributing to the pathogenesis of ischaemia–reperfusion injury. Platelet-activating factor (PAF), a phospholipid mediator of inflammation, has been implicated in the pathogenesis of this process. We previously showed that the infusion of streptokinase (SK) induces the intravascular release of PAF in patients with acute myocardial infarction (AMI), and that cultured human endothelial cells (EC) synthesized PAF in response to SK and plasmin (PLN). In the present study, we investigated the role of the membrane attack complex (MAC) of complement in the PLN-induced synthesis of PAF. In vivo, we showed a correlation between the levels of soluble terminal complement components (sC5b-9) and the concentrations of PAF detected in blood of patients with AMI infused with SK. In vitro both EC and polymorphonuclear neutrophils (PMN), incubated in the presence of PLN and normal human serum, showed an intense staining for the MAC neoepitope, while no staining was detected when they were incubated with PLN in the presence of heat-inactivated normal human serum. Moreover, the insertion of MAC on EC and PMN plasmamembrane elicited the synthesis of PAF. In conclusion, our results elucidate the mechanisms involved in PAF production during the activation of the fibrinolytic system, showing a role for complement products in this setting. The release of PAF may increase the inflammatory response, thus limiting the beneficial effects of thrombolytic therapy. Moreover, it may have a pathogenic role in other pathological conditions, such as transplant rejection, tumoral angiogenesis, and septic shock, where fibrinolysis is activated. PMID:12871223

The membrane attack complex of complement contributes to plasmin-induced synthesis of platelet-activating factor by endothelial cells and neutrophils.

PubMed

Lupia, Enrico; Del Sorbo, Lorenzo; Bergerone, Serena; Emanuelli, Giorgio; Camussi, Giovanni; Montrucchio, Giuseppe

2003-08-01

Thrombolytic agents, used to restore blood flow to ischaemic tissues, activate several enzymatic systems with pro-inflammatory effects, thus potentially contributing to the pathogenesis of ischaemia-reperfusion injury. Platelet-activating factor (PAF), a phospholipid mediator of inflammation, has been implicated in the pathogenesis of this process. We previously showed that the infusion of streptokinase (SK) induces the intravascular release of PAF in patients with acute myocardial infarction (AMI), and that cultured human endothelial cells (EC) synthesized PAF in response to SK and plasmin (PLN). In the present study, we investigated the role of the membrane attack complex (MAC) of complement in the PLN-induced synthesis of PAF. In vivo, we showed a correlation between the levels of soluble terminal complement components (sC5b-9) and the concentrations of PAF detected in blood of patients with AMI infused with SK. In vitro both EC and polymorphonuclear neutrophils (PMN), incubated in the presence of PLN and normal human serum, showed an intense staining for the MAC neoepitope, while no staining was detected when they were incubated with PLN in the presence of heat-inactivated normal human serum. Moreover, the insertion of MAC on EC and PMN plasmamembrane elicited the synthesis of PAF. In conclusion, our results elucidate the mechanisms involved in PAF production during the activation of the fibrinolytic system, showing a role for complement products in this setting. The release of PAF may increase the inflammatory response, thus limiting the beneficial effects of thrombolytic therapy. Moreover, it may have a pathogenic role in other pathological conditions, such as transplant rejection, tumoral angiogenesis, and septic shock, where fibrinolysis is activated.

Expression of a plant-derived peptide harboring water-cleaning and antimicrobial activities.

PubMed

Suarez, M; Entenza, J M; Doerries, C; Meyer, E; Bourquin, L; Sutherland, J; Marison, I; Moreillon, P; Mermod, N

2003-01-05

Drinking water is currently a scarce world resource, the preparation of which requires complex treatments that include clarification of suspended particles and disinfection. Seed extracts of Moringa oleifera Lam., a tropical tree, have been proposed as an environment-friendly alternative, due to their traditional use for the clarification of drinking water. However, the precise nature of the active components of the extract and whether they may be produced in recombinant form are unknown. Here we show that recombinant or synthetic forms of a cationic seed polypeptide mediate efficient sedimentation of suspended mineral particles and bacteria. Unexpectedly, the polypeptide was also found to possesses a bactericidal activity capable of disinfecting heavily contaminated water. Furthermore, the polypeptide has been shown to efficiently kill several pathogenic bacteria, including antibiotic-resistant isolates of Staphylococcus, Streptococcus, and Legionella species. Thus, this polypeptide displays the unprecedented feature of combining water purification and disinfectant properties. Identification of an active principle derived from the seed extracts points to a range of potential for drinking water treatment or skin and mucosal disinfection in clinical settings. Copyright 2002 Wiley Periodicals, Inc.

[The bactericidal effect of oxybuprocaine. A possible cause of false negative results in bronchoalveolar lavage].

PubMed

Anding, K; Albrecht, P; Heilmann, C; Daschner, F

1993-09-01

New approaches in the diagnosis of pneumonia, especially in intensive care units, are quantitative cultures of bronchoalveolar (BAL) fluid or the protected specimen brush. The sensitivity of these methods, however, has often been found to be as low as 50-60%. One possible explanation for the low sensitivity of these diagnostic tools is the antimicrobial activity of local anaesthetics used in bronchoscopy. Therefore, we investigated the bactericidal properties of oxybuprocaine, a topical anaesthetic used for bronchoscopy in our clinic, in order to test the reliability of specimens obtained from BAL. METHODS. The bactericidal activity of oxybuprocaine in concentrations of 1%, 0.1%, 0.05% and 0.01% was tested by constructing time-kill curves for Streptococcus pneumoniae, Hemophilus influenzae, Pseudomonas aeruginosa and Escherichia coli. Five stains of each bacterial species were tested. The inoculum size was 10(4) ml, and bacteria were counted after 10, 20, 30, 60 and 120 min. RESULTS. The resulting time-kill curves are demonstrated in Figs. 1-4. The most sensitive bacteria were S. pneumoniae and H. influenzae, in which significant bactericidal activity could be shown even with a 0.01% solution of oxybuprocaine. E. coli and P. aeruginosa were also inhibited, but only at the highest concentration of 1%. CONCLUSIONS. The use of local anaesthetics before material is taken for culture, e.g. from BAL, may give rise to false-negative results and should therefore be avoided or reduced. For each local anaesthetic used in bronchoscopy, the concentrations that can be used without the risk of false-negative results should be determined.

Bactericidal activity and silver release of porous ceramic candle filter prepared by sintering silica with silver nanoparticles/zeolite for water disinfection

NASA Astrophysics Data System (ADS)

Trinh Nguyen, Thuy Ai; Phu Dang, Van; Duy Nguyen, Ngoc; Le, Anh Quoc; Thanh Nguyen, Duc; Hien Nguyen, Quoc

2014-09-01

Porous ceramic candle filters (PCCF) were prepared by sintering silica from rice husk with silver nanoparticles (AgNPs)/zeolite A at about 1050 °C to create bactericidal PCCF/AgNPs for water disinfection. The silver content in PCCF/AgNPs was of 300-350 mg kg-1 determined by inductively coupled plasma-atomic emission spectroscopy (ICP-AES) and the average pore size of PCCF/AgNPs was of 50-70 Å measured by Brunauer-Emmett-Teller (BET) method. The bactericidal activity and silver release of PCCF/AgNPs have been investigated by flow test with water flow rate of 5 L h-1 and initial inoculation of E. coli in inlet water of 106 CFU/100 mL. The volume of filtrated water was collected up to 500 L. Results showed that the contamination of E. coli in filtrated water was <1 CFU/100 mL and the content of silver released from PCCF/AgNPs into filtrated water was <1 μg L-1, it is low, far under the WHO guideline of 100 μg L-1 at maximum for drinking water. Based on the content of silver in PCCF/AgNPs and in filtrated water, it was estimated that one PCCF/AgNPs could be used to filtrate of ˜100 m3 water. Thus, as-prepared PCCF/AgNPs releases low content of silver into water and shows effectively bactericidal activity that is promising to apply as point-of-use water treatment technology for drinking water disinfection.

Influence of heat inactivation of human serum on the opsonization of Streptococcus mutans.

PubMed

Moore, M A; Hakki, Z W; Gregory, R L; Gfell, L E; Kim-Park, W K; Kowolik, M J

1997-12-15

Phagocytosis of bacteria, such as Streptococcus mutans, is important to host defense. One mechanism by which phagocytosis can be enhanced is by antibody or complement-mediated opsonization of bacteria. Many studies utilize opsonization of bacteria to enhance a cellular response, but little information has been found examining methodology or validity of the opsonization process following the denaturization of the serum. Human serum was inactivated by heat in order to disrupt the classical and alternative pathways of the complement cascade. S. mutans isolated from human subjects were opsonized with heat-inactivated human serum before exposing them to viable neutrophils in vitro. Luminol-dependent chemiluminescence (CL) was used to measure neutrophil activation. Human serum used to opsonize the bacteria was denatured by incubation at 57 degrees C for intervals of 30 and 60 min to inactivate complement. The results from the opsonization data indicated that there was significantly increased CL with 60-min inactivation of the serum (34% increase in mean integration mV.min; p < or = 0.05) over the nonopsonized control. This indicated a successful opsonization of the bacteria. In addition, the data demonstrate that the inactivation of serum requires a minimum of 60 min at 57 degrees C to disrupt the complement cascade, while 30- and 15-min inactivations produced no significant increase in CL activity over the control. Standard sandwich ELISA assays, detecting complement binding to S. mutans, confirmed successful heat inactivation of serum showing a significant decrease (p < or = 0.001) in complement binding to S. mutans after 30 min, but could not explain the increased CL response after 60-min heat deactivation of the serum.

Bactericidal Activity of Ceragenin CSA-13 in Cell Culture and in an Animal Model of Peritoneal Infection.

PubMed

Bucki, Robert; Niemirowicz, Katarzyna; Wnorowska, Urszula; Byfield, Fitzroy J; Piktel, Ewelina; Wątek, Marzena; Janmey, Paul A; Savage, Paul B

2015-10-01

Ceragenins constitute a novel family of cationic antibiotics characterized by a broad spectrum of antimicrobial activities, which have mostly been assessed in vitro. Using a polarized human lung epithelial cell culture system, we evaluated the antibacterial activities of the ceragenin CSA-13 against two strains of Pseudomonas aeruginosa (PAO1 and Xen5). Additionally, the biodistribution and bactericidal activity of a CSA-13-IRDye 800CW derivate were assessed using an animal model of peritoneal infection after PAO1 challenge. In cell culture, CSA-13 bactericidal activities against PAO1 and Xen5 were higher than the activities of the human cathelicidin peptide LL-37. Increased CSA-13 activity was observed in polarized human lung epithelial cell cultures subjected to butyric acid treatment, which is known to increase endogenous LL-37 production. Eight hours after intravenous or intraperitoneal injection, the greatest CSA-13-IRDye 800CW accumulation was observed in mouse liver and kidneys. CSA-13-IRDye 800CW administration resulted in decreased bacterial outgrowth from abdominal fluid collected from animals subjected to intraperitoneal PAO1 infection. These observations indicate that CSA-13 may synergistically interact with antibacterial factors that are naturally present at mucosal surfaces and it maintains its antibacterial activity in the infected abdominal cavity. Cationic lipids such as CSA-13 represent excellent candidates for the development of new antibacterial compounds. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Validation of CoaBC as a Bactericidal Target in the Coenzyme A Pathway of Mycobacterium tuberculosis.

PubMed

Evans, Joanna C; Trujillo, Carolina; Wang, Zhe; Eoh, Hyungjin; Ehrt, Sabine; Schnappinger, Dirk; Boshoff, Helena I M; Rhee, Kyu Y; Barry, Clifton E; Mizrahi, Valerie

2016-12-09

Mycobacterium tuberculosis relies on its own ability to biosynthesize coenzyme A to meet the needs of the myriad enzymatic reactions that depend on this cofactor for activity. As such, the essential pantothenate and coenzyme A biosynthesis pathways have attracted attention as targets for tuberculosis drug development. To identify the optimal step for coenzyme A pathway disruption in M. tuberculosis, we constructed and characterized a panel of conditional knockdown mutants in coenzyme A pathway genes. Here, we report that silencing of coaBC was bactericidal in vitro, whereas silencing of panB, panC, or coaE was bacteriostatic over the same time course. Silencing of coaBC was likewise bactericidal in vivo, whether initiated at infection or during either the acute or chronic stages of infection, confirming that CoaBC is required for M. tuberculosis to grow and persist in mice and arguing against significant CoaBC bypass via transport and assimilation of host-derived pantetheine in this animal model. These results provide convincing genetic validation of CoaBC as a new bactericidal drug target.

Structure-anti-MRSA activity relationship of macrocyclic bis(bibenzyl) derivatives.

PubMed

Sawada, Hiromi; Onoda, Kenji; Morita, Daichi; Ishitsubo, Erika; Matsuno, Kenji; Tokiwa, Hiroaki; Kuroda, Teruo; Miyachi, Hiroyuki

2013-12-15

We synthesized a series of macrocyclic bis(bibenzyl) derivatives, including riccardin-, isoplagiochin- and marchantin-class structures, and evaluated their antibacterial activity towards methicillin-resistant Staphylococcus aureus (anti-MRSA activity). The structure-activity relationships and the results of molecular dynamics simulations indicated that bis(bibenzyl)s with potent anti-MRSA activity commonly have a 4-hydroxyl group at the D-benzene ring and a 2-hydroxyl group at the C-benzene ring in the hydrophilic part of the molecule, and an unsubstituted phenoxyphenyl group in the hydrophobic part of the molecule containing the A-B-benzene rings. Pharmacological characterization of the bis(bibenzyl) derivatives and 2-phenoxyphenol fragment 25, previously proposed as the minimum structure of riccardin C 1 for anti-MRSA activity, indicated that they have different action mechanisms: the bis(bibenzyl)s are bactericidal, while 25 is bacteriostatic, showing only weak bactericidal activity. Copyright © 2013 Elsevier Ltd. All rights reserved.

Cationic Biomimetic Particles of Polystyrene/Cationic Bilayer/Gramicidin for Optimal Bactericidal Activity.

PubMed

Xavier, Gabriel R S; Carmona-Ribeiro, Ana M

2017-12-02

Nanostructured particles of polystyrene sulfate (PSS) covered by a cationic lipid bilayer of dioctadecyldimethylammonium bromide (DODAB) incorporated gramicidin D (Gr) yielding optimal and broadened bactericidal activity against both Escherichia coli and Staphylococcus aureus . The adsorption of DODAB/Gr bilayer onto PSS nanoparticles (NPs) increased the zeta-average diameter by 8-10 nm, changed the zeta-potential of the NPs from negative to positive, and yielded a narrow size distributions for the PSS/DODAB/Gr NPs, which displayed broad and maximal microbicidal activity at very small concentrations of the antimicrobials, namely, 0.057 and 0.0057 mM DODAB and Gr, respectively. The results emphasized the advantages of highly-organized, nanostructured, and cationic particles to achieve hybrid combinations of antimicrobials with broad spectrum activity at considerably reduced DODAB and Gr concentrations.

A versatile synthesis of highly bactericidal Myramistin® stabilized silver nanoparticles

NASA Astrophysics Data System (ADS)

Vertelov, G. K.; Krutyakov, Yu A.; Efremenkova, O. V.; Olenin, A. Yu; Lisichkin, G. V.

2008-09-01

Silver nanoparticles stabilized by a well-known antibacterial surfactant benzyldimethyl[3-(myristoylamino)propyl]ammonium chloride (Myramistin®) were produced for the first time by borohydride reduction of silver chloride sol in water. Stable aqueous dispersions of silver nanoparticles without evident precipitation for several months could be obtained. In vitro bactericidal tests showed that Myramistin® capped silver NPs exhibited notable activity against six different microorganisms—gram-positive and gram-negative bacteria, yeasts and fungi. The activity was up to 20 times higher (against E. coli) compared to Myramistin® at the same concentrations and on average 2 times higher if compared with citrate-stabilized NPs.

Comprehensive bactericidal activity of an ethanol-based hand gel in 15 seconds.

PubMed

Kampf, Günter; Hollingsworth, Angela

2008-01-22

Some studies indicate that the commonly recommended 30 s application time for the post contamination treatment of hands may not be necessary as the same effect may be achieved with some formulations in a shorter application time such as 15 s. We evaluated the bactericidal activity of an ethanol-based hand gel (Sterillium Comfort Gel) within 15 s in a time-kill-test against 11 Gram-positive, 16 Gram-negative bacteria and 11 emerging bacterial pathogens. Each strain was evaluated in quadruplicate. The hand gel (85% ethanol, w/w) was found to reduce all 11 Gram-positive and all 16 Gram-negative bacteria by more than 5 log10 steps within 15 s, not only against the ATCC test strains but also against corresponding clinical isolates. In addition, a log10 reduction > 5 was observed against all tested emerging bacterial pathogens. The ethanol-based hand gel was found to have a broad spectrum of bactericidal activity in only 15 s which includes the most common species causing nosocomial infections and the relevant emerging pathogens. Future research will hopefully help to find out if a shorter application time for the post contamination treatment of hands provides more benefits or more risks.