scholarly journals Participation of a galactose-specific C-type lectin in Drosophila immunity

2006 ◽  
Vol 396 (1) ◽  
pp. 127-138 ◽  
Author(s):  
Takahiro Tanji ◽  
Ayako Ohashi-Kobayashi ◽  
Shunji Natori
Keyword(s):  
Fruit Fly ◽  
Biological Properties ◽  
Erwinia Chrysanthemi ◽  
Antibacterial Peptides ◽  
Null Mutant ◽  
Gram Negative ◽  
E Coli ◽  
Microbial Strains ◽  
Lectin Genes

A galactose-specific C-type lectin has been purified from a pupal extract of Drosophila melanogaster. This lectin gene, named DL1 (Drosophilalectin 1), is part of a gene cluster with the other two galactose-specific C-type lectin genes, named DL2 (Drosophilalectin 2) and DL3 (Drosophilalectin 3). These three genes are expressed differentially in fruit fly, but show similar haemagglutinating activities. The present study characterized the biochemical and biological properties of the DL1 protein. The recombinant DL1 protein bound to Escherichia coli and Erwinia chrysanthemi, but not to other Gram-negative or any other kinds of microbial strains that have been investigated. In addition, DL1 agglutinated E. coli and markedly intensified the association of a Drosophila haemocytes-derived cell line with E. coli. For in vivo genetic analysis of the lectin genes, we also established a null-mutant Drosophila. The induction of inducible antibacterial peptide genes was not impaired in the DL1 mutant, suggesting that the galactose-specific C-type lectin does not participate in the induction of antibacterial peptides, but possibly participates in the immune response via the haemocyte-mediated mechanism.

scholarly journals In Vitro and In Vivo Antibacterial Activities of a Novel Glycylcycline, the 9-t-Butylglycylamido Derivative of Minocycline (GAR-936)

1999 ◽  
Vol 43 (4) ◽  
pp. 738-744 ◽  
Author(s):  
P. J. Petersen ◽  
N. V. Jacobus ◽  
W. J. Weiss ◽  
P. E. Sum ◽  
R. T. Testa
Keyword(s):  
Anaerobic Bacteria ◽  
Tetracycline Resistance ◽  
Protective Effects ◽  
Gram Negative ◽  
E Coli ◽  
Resistance Determinants ◽  
Aerobic And Anaerobic ◽  
Ribosomal Protection

ABSTRACT The 9-t-butylglycylamido derivative of minocycline (TBG-MINO) is a recently synthesized member of a novel group of antibiotics, the glycylcyclines. This new derivative, like the first glycylcyclines, theN,N-dimethylglycylamido derivative of minocycline and 6-demethyl-6-deoxytetracycline, possesses activity against bacterial isolates containing the two major determinants responsible for tetracycline resistance: ribosomal protection and active efflux. The in vitro activities of TBG-MINO and the comparative agents were evaluated against strains with characterized tetracycline resistance as well as a spectrum of recent clinical aerobic and anaerobic gram-positive and gram-negative bacteria. TBG-MINO, with an MIC range of 0.25 to 0.5 μg/ml, showed good activity against strains expressing tet(M) (ribosomal protection), tet(A), tet(B),tet(C), tet(D), and tet(K) (efflux resistance determinants). TBG-MINO exhibited similar activity against methicillin-resistant Staphylococcus aureus (MRSA), penicillin-resistant streptococci, and vancomycin-resistant enterococci (MICs at which 90% of strains are inhibited, ≤0.5 μg/ml). TBG-MINO exhibited activity against a wide diversity of gram-negative aerobic and anaerobic bacteria, most of which were less susceptible to tetracycline and minocycline. The in vivo protective effects of TBG-MINO were examined against acute lethal infections in mice caused by Escherichia coli, S. aureus, andStreptococcus pneumoniae isolates. TBG-MINO, administered intravenously, demonstrated efficacy against infections caused byS. aureus including MRSA strains and strains containingtet(K) or tet(M) resistance determinants (median effective doses [ED50s], 0.79 to 2.3 mg/kg of body weight). TBG-MINO demonstrated efficacy against infections caused by tetracycline-sensitive E. coli strains as well asE. coli strains containing either tet(M) or the efflux determinant tet(A), tet(B), ortet(C) (ED50s, 1.5 to 3.5 mg/kg). Overall, TBG-MINO shows antibacterial activity against a wide spectrum of gram-positive and gram-negative aerobic and anaerobic bacteria including strains resistant to other chemotherapeutic agents. The in vivo protective effects, especially against infections caused by resistant bacteria, corresponded with the in vitro activity of TBG-MINO.

scholarly journals Non-Lytic Antibacterial Peptides That Translocate Through Bacterial Membranes to Act on Intracellular Targets

2019 ◽  
Vol 20 (19) ◽  
pp. 4877 ◽  
Author(s):  
Marlon H. Cardoso ◽  
Beatriz T. Meneguetti ◽  
Bruna O. Costa ◽  
Danieli F. Buccini ◽  
Karen G. N. Oshiro ◽  
...  
Keyword(s):  
Cell Wall ◽  
Protein Synthesis ◽  
Pathogenic Bacteria ◽  
Biological Properties ◽  
Antibacterial Peptides ◽  
Bacterial Cells ◽  
Bacterial Membranes ◽  
Intracellular Targets

The advent of multidrug resistance among pathogenic bacteria has attracted great attention worldwide. As a response to this growing challenge, diverse studies have focused on the development of novel anti-infective therapies, including antimicrobial peptides (AMPs). The biological properties of this class of antimicrobials have been thoroughly investigated, and membranolytic activities are the most reported mechanisms by which AMPs kill bacteria. Nevertheless, an increasing number of works have pointed to a different direction, in which AMPs are seen to be capable of displaying non-lytic modes of action by internalizing bacterial cells. In this context, this review focused on the description of the in vitro and in vivo antibacterial and antibiofilm activities of non-lytic AMPs, including indolicidin, buforin II PR-39, bactenecins, apidaecin, and drosocin, also shedding light on how AMPs interact with and further translocate through bacterial membranes to act on intracellular targets, including DNA, RNA, cell wall and protein synthesis.

scholarly journals 1364. Efficacy of Omadacycline Against Molecularly Characterized Gram-Positive and Gram-Negative Pathogens Causing Infections in the Phase 3 CABP and ABSSSI Clinical Trials

2018 ◽  
Vol 5 (suppl_1) ◽  
pp. S417-S418 ◽  
Author(s):  
Rodrigo E Mendes ◽  
Mariana Castanheira ◽  
Eliana S Armstrong ◽  
Judith N Steenbergen ◽  
Robert K Flamm
Keyword(s):  
Efflux Pump ◽  
Clinical Success ◽  
Treated Patient ◽  
Research Support ◽  
Phase 3 ◽  
Gram Positive ◽  
Two Phase ◽  
Gram Negative ◽  
E Coli

Abstract Background Omadacycline (OMC) is a once-daily agent with IV and oral formulations. One Phase 3 trial for community-acquired bacterial pneumonia (CABP; OPTIC) and two Phase 3 trials for acute bacterial skin and skin structure infections (ABSSSI; OASIS-1 and OASIS-2) were completed. OMC and comparator efficacies were examined for molecularly characterized baseline pathogens. Methods Gram-positive (24) and -negative (17) isolates from the OPTIC (26), OASIS-1 (10) or OASIS-2 (5) trials were selected for characterization. Susceptibility testing and interpretation was performed by CLSI methods. Gram-positive isolates were selected based on tetracycline and/or macrolide, lincosamide, streptograminB (MLSB) phenotypes, and tetracycline-nonsusceptible (NS) Gram-negative isolates were selected. Isolates were subjected to next-generation sequencing followed by screening for known tetracycline and/or MLSB genes. The efficacy endpoint was investigator’s assessment of clinical response at post therapy evaluation (PTE). Results All S. aureus (eight isolates) exhibited a doxycycline-NS phenotype (MIC, 8–16 µg/mL) and OMC MIC values of 0.25–0.5 µg/mL. All S. aureus carried tet(M), except one isolate with tet(K), and one isolate with tet(M) and tet(L). All but one S. pneumoniae (16 isolates; OMC MIC, 0.03–0.06 µg/mL) carried MLSB genes, while tetracycline- and doxycycline-NS isolates (12) had tet(M). E. coli (eight isolates; OMC MIC, 0.5–2 µg/mL), E. cloacae (two isolates; OMC MIC, 2 µg/mL), and K. pneumoniae (six isolates; OMC MIC, 2–16 µg/mL) carried tetracycline efflux-pump genes, tet(A) and/or tet(B), tet(D), and tet(A), respectively. tet genes were not detected in one K. pneumoniae (OMC MIC, 8 µg/mL). Clinical success was noted in 37/41 (90.2%) patients. Two linezolid-treated patients with S. aureus (OMC MIC, 0.25 µg/mL) from OASIS-1 and one OMC-treated patient from OPTIC with E. coli (OMC MIC, 2 µg/mL) had indeterminate PTE responses. One OMC-treated patient from OPTIC with K. pneumoniae (OMC MIC, 8 µg/mL) was a clinical failure at PTE. Conclusion This study expands on OMC efficacy data analysis among patients infected with tetracycline–NS pathogens. These results indicate that OMC in vivo efficacy is not affected by tetracycline and/or MLSB resistance mechanisms. Disclosures R. E. Mendes, Paratek Pharmaceuticals: Research Contractor, Research support. M. Castanheira, Paratek Pharmaceuticals: Research Contractor, Research support. E. S. Armstrong, Paratek Pharmaceuticals: Employee, Salary. J. N. Steenbergen, Paratek Pharmaceuticals: Employee and Shareholder, Salary. R. K. Flamm, Paratek Pharmaceuticals: Scientific Advisor, Speaker honorarium.

scholarly journals Full Capacity of Recombinant Escherichia coliHeat-Stable Enterotoxin Fusion Proteins for Extracellular Secretion, Antigenicity, Disulfide Bond Formation, and Activity

2000 ◽  
Vol 68 (7) ◽  
pp. 4064-4074 ◽  
Author(s):  
Isabelle Batisson ◽  
Maurice Der Vartanian ◽  
Brigitte Gaillard-Martinie ◽  
Michel Contrepois
Keyword(s):  
Disulfide Bonds ◽  
Secretory Pathway ◽  
Biological Properties ◽  
Leader Peptide ◽  
Dependent Manner ◽  
Reporter Protein ◽  
E Coli ◽  
Heat Stable

ABSTRACT We have successfully used the major subunit ClpG ofEscherichia coli CS31A fimbriae as an antigenic and immunogenic exposure-delivery vector for various heterologous peptides varying in nature and length. However, the ability of ClpG as a carrier to maintain in vitro and in vivo the native biological properties of passenger peptide has not yet been reported. To address this possibility, we genetically fused peptides containing all or part of the E. coli human heat-stable enterotoxin (STh) sequence to the amino or carboxyl ends of ClpG. Using antibodies to the ClpG and STh portions for detecting the hybrids; AMS (4-acetamido-4′-maleimidylstilbene-2,2′-disulfonate), a potent free thiol-trapping reagent, for determining the redox state of STh in the fusion; and the suckling mouse assay for enterotoxicity, we demonstrated that all ClpG-STh proteins were secreted in vitro and in vivo outside the E. coli cells in a heat-stable active oxidized (disulfide-bonded) form. Indeed, in contrast to many earlier studies, blocking the natural NH2 or COOH extremities of STh had, in all cases, no drastic effect on cell release and toxin activity. Only antigenicity of STh C-terminally extended with ClpG was strongly affected in a conformation-dependent manner. These results suggest that the STh activity was not altered by the chimeric structure, and therefore that, like the natural toxin, STh in the fusion had a spatial structure flexible enough to be compatible with secretion and enterotoxicity (folding and STh receptor recognition). Our study also indicates that disulfide bonds were essential for enterotoxicity but not for release, that spontaneous oxidation by molecular oxygen occurred in vitro in the medium, and that the E. coli cell-bound toxin activity in vivo resulted from an effective export processing of hybrids and not cell lysis. None of the ClpG-STh subunits formed hybrid CS31A-STh fimbriae at the cell surface ofE. coli, and a strong decrease in the toxin activity was observed in the absence of CS31A helper proteins. In fact, chimeras translocated across the outer membrane as a free folded monomer once they were guided into the periplasm by the ClpG leader peptide through the CS31A-dependent secretory pathway. In summary, ClpG appears highly attractive as a carrier reporter protein for basic and applied research through the engineering of E. coli for culture supernatant delivery of an active cysteine-containing protein, such as the heat-stable enterotoxin.

scholarly journals Novel Leu-Val Based Dipeptide as Antimicrobial and Antimalarial Agents: Synthesis and Molecular Docking

2020 ◽  
Vol 8 ◽  
Author(s):  
James A. Ezugwu ◽  
Uchechukwu C. Okoro ◽  
Mercy A. Ezeokonkwo ◽  
China R. Bhimapaka ◽  
Sunday N. Okafor ◽  
...  
Keyword(s):  
Antioxidant Activities ◽  
Condensation Reaction ◽  
Biological Properties ◽  
Health Issues ◽  
Protein Residues ◽  
Antimalarial Agents ◽  
Antimalarial Resistance ◽  
Microbial Strains

The increase of antimicrobial resistance (AMR) and antimalarial resistance are complex and severe health issues today, as many microbial strains have become resistant to market drugs. The choice for the synthesis of new dipeptide-carboxamide derivatives is as a result of their wide biological properties such as antimicrobial, anti-inflammatory, and antioxidant activities. The condensation reaction of substituted benzenesulphonamoyl pentanamides with the carboxamide derivatives using peptide coupling reagents gave targeted products (8a-j). The in silico antimalarial and antibacterial studies showed good interactions of the compounds with target protein residues and a higher dock score in comparison with standard drugs. In the in vivo study, compound 8j was the most potent antimalarial agent with 61.90% inhibition comparable with 67% inhibition for Artemisinin. In the in vitro antimicrobial activity, compounds 8a and 8b (MIC 1.2 × 10−3 M and 1.1 × 10−3 M) were most potent against S. aureus; compound 8a, 8b, and 8j with MIC 6.0 × 10−3 M, 5.7 × 10−4 M, and 6.5 × 10−4 M, respectively, were the most active against B. subtilis; compound 8b (MIC 9.5 × 10−4 M) was most active against E.coli while 8a, 8b and 8d were the most active against S. typhi. Compounds 8c and 8h (MIC 1.3 × 10−3 M) each were the most active against C. albicans, while compound 8b (MIC 1.3 × 10−4 M) was most active against A. niger.

scholarly journals Antimicrobial Drug Resistance Genes Do Not Convey a Secondary Fitness Advantage to Calf-Adapted Escherichia coli

2006 ◽  
Vol 72 (1) ◽  
pp. 443-448 ◽  
Author(s):  
Artashes R. Khachatryan ◽  
Dale D. Hancock ◽  
Thomas E. Besser ◽  
Douglas R. Call
Keyword(s):  
Antimicrobial Resistance ◽  
Resistance Genes ◽  
Null Mutant ◽  
Antimicrobial Drug Resistance ◽  
E Coli ◽  
Fitness Advantage ◽  
Antimicrobial Resistance Genes ◽  
Mutant Strains

ABSTRACT Maintenance of antimicrobial drug resistance in bacteria can be influenced by factors unrelated to direct selection pressure such as close linkage to other selectively advantageous genes and secondary advantage conveyed by antimicrobial resistance genes in the absence of drug selection. Our previous trials at a dairy showed that the maintenance of the antimicrobial resistance genes is not influenced by specific antimicrobial selection and that the most prevalent antimicrobial resistance phenotype of Escherichia coli is specifically selected for in young calves. In this paper we examine the role of secondary advantages conveyed by antimicrobial resistance genes. We tested antimicrobial-susceptible null mutant strains for their ability to compete with their progenitor strains in vitro and in vivo. The null mutant strains were generated by selection for spontaneous loss of resistance genes in broth supplemented with fusaric acid or nickel chloride. On average, the null mutant strains were as competitive as the progenitor strains in vitro and in newborn calves (in vivo). Inoculation of newborn calves at the dairy with antimicrobial-susceptible strains of E. coli did not impact the prevalence of antimicrobial-resistant E. coli. Our results demonstrate that the antimicrobial resistance genes are not responsible for the greater fitness advantage of antimicrobial-resistant E. coli in calves, but the farm environment and the diet clearly exert critical selective pressures responsible for the maintenance of antimicrobial resistance genes. Our current hypothesis is that the antimicrobial resistance genes are linked to other genes responsible for differential fitness in dairy calves.

scholarly journals Degradation of epigallocatechin and epicatechin gallates by a novel tannase TanHcw from Herbaspirillum camelliae

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Jia Lei ◽  
Yong Zhang ◽  
Xuechen Ni ◽  
Xuejing Yu ◽  
Xingguo Wang
Keyword(s):  
Enzyme Activity ◽  
Biological Properties ◽  
Tea Plant ◽  
Optimal Temperature ◽  
Gram Negative ◽  
E Coli ◽  
Its Gene ◽  
N Terminus ◽  
Tea Plants ◽  
Inhibited Enzyme

Abstract Background Herbaspirillum camelliae is a gram-negative endophyte isolated from the tea plant. Both strains WT00C and WT00F were found to hydrolyze epigallocatechin-3-gallate (EGCG) and epicatechin-3-gallate (ECG) to release gallic acid (GA) and display tannase activity. However, no tannase gene was annotated in the genome of H. camelliae WT00C. Results The 39 kDa protein, annotated as the prolyl oligopeptidase in the NCBI database, was finally identified as a novel tannase. Its gene was cloned, and the enzyme was expressed in E. coli and purified to homogeneity. Moreover, enzymatic characterizations of this novel tannase named TanHcw were studied. TanHcw was a secretary enzyme with a Sec/SPI signal peptide of 48 amino acids at the N-terminus, and it catalyzed the degradation of tannin, methyl gallate (MG), epigallocatechin-3-gallate (EGCG) and epicatechin-3-gallate (ECG). The optimal temperature and pH of TanHcw activities were 30 °C, pH 6.0 for MG and 40 °C, pH 7.0 for both EGCG and ECG. Na+, K+ Mn2+ and Triton-X100, Tween80 increased the enzyme activity of TanHcw, whereas Zn2+, Mg2+, Hg2+, EMSO, EDTA and β-mercaptoethanol inhibited enzyme activity. Km, kcat and kcat /Km of TanHcw were 0.30 mM, 37.84 s−1, 130.67 mM−1 s−1 for EGCG, 0.33 mM, 34.59 s−1, 105.01 mM−1 s−1 for ECG and 0.82 mM, 14.64 s−1, 18.17 mM−1 s−1 for MG, respectively. Conclusion A novel tannase TanHcw from H. camelliae has been identified and characterized. The biological properties of TanHcw suggest that it plays a crucial role in the specific colonization of H. camelliae in tea plants. Discovery of the tannase TanHcw in this study gives us a reasonable explanation for the host specificity of H. camelliae. In addition, studying the characteristics of this enzyme offers the possibility of further defining its potential in industrial application.

scholarly journals Hybrid Approach for Synthesis and Antimicrobial Activity of Heterocyclic Compounds

2018 ◽  
Vol 77 ◽  
pp. 35-52 ◽  
Author(s):  
Nisheeth C. Desai ◽  
Darshita V. Vaja
Keyword(s):  
Antimicrobial Activity ◽  
Heterocyclic Compounds ◽  
Hybrid Approach ◽  
Spectroscopic Techniques ◽  
Gram Negative Bacteria ◽  
Gram Positive ◽  
Gram Negative ◽  
E Coli ◽  
Microbial Strains

We have synthesized novel series of N-(1-(2-(1-phenyl-3-(p-tolyl)-1H-pyrazol-4-yl)-5-(pyridin-4-yl)-1,3,4-oxadiazol-3(2H)-yl)ethylidene)arylaniline and their derivatives. The structures of synthesized compounds were well characterized by spectroscopic techniques. Antimicrobial activity of the newly synthesized derivatives was evaluated against gram positive (S. aureus and S. pyogenes), gram negative bacteria (E. coli and P. aeruginosa), and strains of fungi (C. albicans, A. niger and A. clavatus). Among the screened derivatives 5c, 5f, 5i, 5l and 5t demonstrated superior antimicrobial activity against microbial strains.

scholarly journals Pharmacodynamic Evaluation of MRX-8, a Novel Polymyxin, in the Neutropenic Mouse Thigh and Lung Infection Models against Gram-Negative Pathogens

2020 ◽  
Vol 64 (11) ◽  
Author(s):  
Alexander J. Lepak ◽  
Wen Wang ◽  
David R. Andes
Keyword(s):  
Polymyxin B ◽  
Lung Model ◽  
Dose Fractionation ◽  
Time Curve ◽  
Gram Negative ◽  
Content Type ◽  
E Coli ◽  
Infection Models ◽  
Dose Ranging

ABSTRACT MRX-8 is a novel polymyxin analogue in development for the treatment of infections caused by Gram-negative pathogens, including those resistant to other antibiotic classes. In the present study, we examined the pharmacodynamic activity of MRX-8 against a variety of common Gram-negative pathogens in the neutropenic mouse thigh and lung models. Additionally, we examined polymyxin B (PMB) as a comparator. Plasma pharmacokinetics of MRX-8 and PMB were linear over a broad dosing range of 0.156 to 10 mg/kg of body weight and had similar AUC0–∞ (area under the drug concentration-time curve from 0 h to infinity) exposures of MRX-8, 0.22 to 12.64 mg · h/liter, and PMB, 0.12 to 13.22 mg · h/liter. Dose fractionation was performed for MRX-8 using a single Escherichia coli isolate, and the results demonstrated that both Cmax (maximum concentration of drug in serum)/MIC and AUC/MIC ratios were strongly associated with efficacy. In the thigh model, dose-ranging studies included strains of E. coli (n = 3), Pseudomonas aeruginosa (n = 2), Klebsiella pneumoniae (n = 3), and Acinetobacter baumannii (n = 1). Both MRX-8 and PMB exhibited increased effects with increasing doses. MRX-8 and PMB free AUC/MIC exposures for net stasis were similar for E. coli and K. pneumoniae at 20 to 30. Notably, for P. aeruginosa and A. baumannii, the free AUC/MIC ratio for stasis was numerically much smaller for MRX-8 at 6 to 8 than for PMB at 16 to 37. In the lung model, MRX-8 was also more effective than PMB when dosed to achieve similar free-drug AUC exposures over the study period. MRX-8 is a promising novel polymyxin analogue with in vivo activity against many different clinically relevant species in both the mouse thigh and lung models.

scholarly journals Conserved mechanism of cell-wall synthase regulation revealed by the identification of a new PBP activator inPseudomonas aeruginosa

2018 ◽  
Vol 115 (12) ◽  
pp. 3150-3155 ◽  
Author(s):  
Neil G. Greene ◽  
Coralie Fumeaux ◽  
Thomas G. Bernhardt
Keyword(s):  
Cell Wall ◽  
Drug Targets ◽  
Bacterial Cells ◽  
Acinetobacter Baylyi ◽  
Gram Negative ◽  
E Coli ◽  
Outer Membrane Lipoprotein ◽  
Synthase Regulation

Penicillin-binding proteins (PBPs) are synthases required to build the essential peptidoglycan (PG) cell wall surrounding most bacterial cells. The mechanisms regulating the activity of these enzymes to control PG synthesis remain surprisingly poorly defined given their status as key antibiotic targets. Several years ago, the outer-membrane lipoproteinEcLpoB was identified as a critical activator ofEscherichia coliPBP1b (EcPBP1b), one of the major PG synthases of this organism. Activation ofEcPBP1b is mediated through the association ofEcLpoB with a regulatory domain onEcPBP1b called UB2H. Notably,Pseudomonas aeruginosaalso encodes PBP1b (PaPBP1b), which possesses a UB2H domain, but this bacterium lacks an identifiable LpoB homolog. We therefore searched for potentialPaPBP1b activators and identified a lipoprotein unrelated to LpoB that is required for the in vivo activity ofPaPBP1b. We named this protein LpoP and found that it interacts directly withPaPBP1b in vitro and is conserved in many Gram-negative species. Importantly, we also demonstrated thatPaLpoP-PaPBP1b as well as an equivalent protein pair fromAcinetobacter baylyican fully substitute forEcLpoB-EcPBP1b inE. colifor PG synthesis. Furthermore, we show that amino acid changes inPaPBP1b that bypass thePaLpoP requirement map to similar locations in the protein as changes promotingEcLpoB bypass inEcPBP1b. Overall, our results indicate that, although different Gram-negative bacteria activate their PBP1b synthases with distinct lipoproteins, they stimulate the activity of these important drug targets using a conserved mechanism.

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