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Gordon L Archer - One of the best experts on this subject based on the ideXlab platform.
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combinations of Lysostaphin with β lactams are synergistic against oxacillin resistant staphylococcus epidermidis
Antimicrobial Agents and Chemotherapy, 2002Co-Authors: Nandini Kiri, Gordon L Archer, Michael W ClimoAbstract:Oxacillin-resistant Staphylococcus aureus is rapidly killed by the endopeptidase Lysostaphin, and the addition of β-lactam antibiotics provides synergistic killing. We investigated the possibility that β-lactams given in combination with Lysostaphin would improve the activity of Lysostaphin against oxacillin-resistant Staphylococcus epidermidis (ORSE), which is normally less susceptible to Lysostaphin. Checkerboard synergy testing was performed for Lysostaphin given in combination with oxacillin against 10 ORSE isolates for which the Lysostaphin MICs were ≥ 8 μg/ml. The fractional inhibitory concentration index ranged from 0.0234 to 0.2656, indicating synergy, which was confirmed in growth curve experiments. In the rabbit model of experimental aortic valve endocarditis using an ORSE strain, the combination of Lysostaphin and nafcillin was as effective as vancomycin alone and significantly better than Lysostaphin or nafcillin alone. We conclude that β-lactam antibiotics given in combination with Lysostaphin are synergistic against many strains of ORSE.
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mechanism and suppression of Lysostaphin resistance in oxacillin resistant staphylococcus aureus
Antimicrobial Agents and Chemotherapy, 2001Co-Authors: Michael W Climo, Kerstin Ehlert, Gordon L ArcherAbstract:The potential for the development of resistance in oxacillin-resistant Staphylococcus aureus (ORSA) to Lysostaphin, a glycylglycine endopeptidase produced by Staphylococcus simulans biovar staphylolyticus, was examined in vitro and in an in vivo model of infection. Following in vitro exposure of ORSA to subinhibitory concentrations of Lysostaphin, Lysostaphin-resistant mutants were idenitifed among all isolates examined. Resistance to Lysostaphin was associated with a loss of resistance to β-lactams and a change in the muropeptide interpeptide cross bridge from pentaglycine to a single glycine. Mutations in femA, the gene required for incorporation of the second and third glycines into the cross bridge, were found following PCR amplification and nucleotide sequence analysis. Complementation of Lysostaphin-resistant mutants with pBBB31, which encodes femA, restored the phenotype of oxacillin resistance and Lysostaphin susceptibility. Addition of β-lactam antibiotics to Lysostaphin in vitro prevented the development of Lysostaphin-resistant mutants. In the rabbit model of experimental endocarditis, administration of a low dose of Lysostaphin for 3 days led predictably to the appearance of Lysostaphin-resistant ORSA mutants in vegetations. Coadministration of nafcillin with Lysostaphin prevented the emergence of Lysostaphin-resistant mutants and led to a mean reduction in aortic valve vegetation counts of 7.5 log10 CFU/g compared to those for untreated controls and eliminated the isolation of Lysostaphin-resistant mutants from aortic valve vegetations. Treatment with nafcillin and Lysostaphin given alone led to mean reductions of 1.35 and 1.65 log10 CFU/g respectively. In ORSA, resistance to Lysostaphin was associated with mutations in femA, but resistance could be suppressed by the coadministration of β-lactam antibiotics.
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Lysostaphin treatment of experimental aortic valve endocarditis caused by a staphylococcus aureus isolate with reduced susceptibility to vancomycin
Antimicrobial Agents and Chemotherapy, 1999Co-Authors: Roberto L Patron, Michael W Climo, Beth P Goldstein, Gordon L ArcherAbstract:The rabbit model of endocarditis was used to test the effectiveness of vancomycin and two different Lysostaphin dosing regimens for the treatment of infections caused by a Staphylococcus aureus strain with reduced susceptibility to vancomycin (glycopeptide-intermediate susceptible S. aureus [GISA]). Vancomycin was ineffective, with no evidence of sterilization of aortic valve vegetations. However, rates of sterilization of aortic valve vegetations were significantly better for animals treated with either a single dose of Lysostaphin (43%) or Lysostaphin given twice daily for 3 days (83%) than for animals treated with vancomycin. Rabbits given a single dose of Lysostaphin followed by a 3-day drug-free period had mean reductions in aortic valve vegetation bacterial counts of 7.27 and 6.63 log 10 CFU/g compared with those for untreated control rabbits and the vancomycin-treated group, respectively. We conclude that Lysostaphin is an effective alternative for the treatment of experimental aortic valve endocarditis caused by a clinical VISA strain.
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Lysostaphin treatment of experimental methicillin resistant staphylococcus aureus aortic valve endocarditis
Antimicrobial Agents and Chemotherapy, 1998Co-Authors: Michael W Climo, Roberto L Patron, Beth P Goldstein, Gordon L ArcherAbstract:The emergence of clinical isolates of methicillin-resistant Staphylococcus aureus with reduced susceptibility to vancomycin has prompted a search for new and novel therapeutic agents active against S. aureus . Lysostaphin, a peptidase produced by Staphylococcus simulans , specifically cleaves the glycine-glycine bonds unique to the interpeptide cross-bridge of the S. aureus cell wall. The effectiveness of various regimens of dosing with intravenous Lysostaphin was compared to that of vancomycin in the rabbit model of aortic valve endocarditis caused by a clinical methicillin-resistant S. aureus isolate. All animals were treated for a total of 3 days. The most active regimen, Lysostaphin given three times daily, produced sterile vegetations in 10 of 11 treated rabbits, with a mean reduction in vegetation bacterial counts of 8.5 log 10 CFU/g compared to the counts in the untreated controls. In contrast, vancomycin given twice daily sterilized no vegetations and reduced vegetation bacterial counts by only 4.8 log 10 CFU/g. Lysostaphin given once daily was less effective, reducing mean vegetation bacterial counts by only 3.6 log 10 CFU/g, but the combination of Lysostaphin once daily and vancomycin twice daily reduced the mean vegetation bacterial density by 7.5 log 10 CFU/g, a result that was significantly better than that for either regimen alone ( P S. aureus aortic valve endocarditis than vancomycin alone.
Michael W Climo - One of the best experts on this subject based on the ideXlab platform.
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combinations of Lysostaphin with β lactams are synergistic against oxacillin resistant staphylococcus epidermidis
Antimicrobial Agents and Chemotherapy, 2002Co-Authors: Nandini Kiri, Gordon L Archer, Michael W ClimoAbstract:Oxacillin-resistant Staphylococcus aureus is rapidly killed by the endopeptidase Lysostaphin, and the addition of β-lactam antibiotics provides synergistic killing. We investigated the possibility that β-lactams given in combination with Lysostaphin would improve the activity of Lysostaphin against oxacillin-resistant Staphylococcus epidermidis (ORSE), which is normally less susceptible to Lysostaphin. Checkerboard synergy testing was performed for Lysostaphin given in combination with oxacillin against 10 ORSE isolates for which the Lysostaphin MICs were ≥ 8 μg/ml. The fractional inhibitory concentration index ranged from 0.0234 to 0.2656, indicating synergy, which was confirmed in growth curve experiments. In the rabbit model of experimental aortic valve endocarditis using an ORSE strain, the combination of Lysostaphin and nafcillin was as effective as vancomycin alone and significantly better than Lysostaphin or nafcillin alone. We conclude that β-lactam antibiotics given in combination with Lysostaphin are synergistic against many strains of ORSE.
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mechanism and suppression of Lysostaphin resistance in oxacillin resistant staphylococcus aureus
Antimicrobial Agents and Chemotherapy, 2001Co-Authors: Michael W Climo, Kerstin Ehlert, Gordon L ArcherAbstract:The potential for the development of resistance in oxacillin-resistant Staphylococcus aureus (ORSA) to Lysostaphin, a glycylglycine endopeptidase produced by Staphylococcus simulans biovar staphylolyticus, was examined in vitro and in an in vivo model of infection. Following in vitro exposure of ORSA to subinhibitory concentrations of Lysostaphin, Lysostaphin-resistant mutants were idenitifed among all isolates examined. Resistance to Lysostaphin was associated with a loss of resistance to β-lactams and a change in the muropeptide interpeptide cross bridge from pentaglycine to a single glycine. Mutations in femA, the gene required for incorporation of the second and third glycines into the cross bridge, were found following PCR amplification and nucleotide sequence analysis. Complementation of Lysostaphin-resistant mutants with pBBB31, which encodes femA, restored the phenotype of oxacillin resistance and Lysostaphin susceptibility. Addition of β-lactam antibiotics to Lysostaphin in vitro prevented the development of Lysostaphin-resistant mutants. In the rabbit model of experimental endocarditis, administration of a low dose of Lysostaphin for 3 days led predictably to the appearance of Lysostaphin-resistant ORSA mutants in vegetations. Coadministration of nafcillin with Lysostaphin prevented the emergence of Lysostaphin-resistant mutants and led to a mean reduction in aortic valve vegetation counts of 7.5 log10 CFU/g compared to those for untreated controls and eliminated the isolation of Lysostaphin-resistant mutants from aortic valve vegetations. Treatment with nafcillin and Lysostaphin given alone led to mean reductions of 1.35 and 1.65 log10 CFU/g respectively. In ORSA, resistance to Lysostaphin was associated with mutations in femA, but resistance could be suppressed by the coadministration of β-lactam antibiotics.
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Lysostaphin treatment of experimental aortic valve endocarditis caused by a staphylococcus aureus isolate with reduced susceptibility to vancomycin
Antimicrobial Agents and Chemotherapy, 1999Co-Authors: Roberto L Patron, Michael W Climo, Beth P Goldstein, Gordon L ArcherAbstract:The rabbit model of endocarditis was used to test the effectiveness of vancomycin and two different Lysostaphin dosing regimens for the treatment of infections caused by a Staphylococcus aureus strain with reduced susceptibility to vancomycin (glycopeptide-intermediate susceptible S. aureus [GISA]). Vancomycin was ineffective, with no evidence of sterilization of aortic valve vegetations. However, rates of sterilization of aortic valve vegetations were significantly better for animals treated with either a single dose of Lysostaphin (43%) or Lysostaphin given twice daily for 3 days (83%) than for animals treated with vancomycin. Rabbits given a single dose of Lysostaphin followed by a 3-day drug-free period had mean reductions in aortic valve vegetation bacterial counts of 7.27 and 6.63 log 10 CFU/g compared with those for untreated control rabbits and the vancomycin-treated group, respectively. We conclude that Lysostaphin is an effective alternative for the treatment of experimental aortic valve endocarditis caused by a clinical VISA strain.
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Lysostaphin treatment of experimental methicillin resistant staphylococcus aureus aortic valve endocarditis
Antimicrobial Agents and Chemotherapy, 1998Co-Authors: Michael W Climo, Roberto L Patron, Beth P Goldstein, Gordon L ArcherAbstract:The emergence of clinical isolates of methicillin-resistant Staphylococcus aureus with reduced susceptibility to vancomycin has prompted a search for new and novel therapeutic agents active against S. aureus . Lysostaphin, a peptidase produced by Staphylococcus simulans , specifically cleaves the glycine-glycine bonds unique to the interpeptide cross-bridge of the S. aureus cell wall. The effectiveness of various regimens of dosing with intravenous Lysostaphin was compared to that of vancomycin in the rabbit model of aortic valve endocarditis caused by a clinical methicillin-resistant S. aureus isolate. All animals were treated for a total of 3 days. The most active regimen, Lysostaphin given three times daily, produced sterile vegetations in 10 of 11 treated rabbits, with a mean reduction in vegetation bacterial counts of 8.5 log 10 CFU/g compared to the counts in the untreated controls. In contrast, vancomycin given twice daily sterilized no vegetations and reduced vegetation bacterial counts by only 4.8 log 10 CFU/g. Lysostaphin given once daily was less effective, reducing mean vegetation bacterial counts by only 3.6 log 10 CFU/g, but the combination of Lysostaphin once daily and vancomycin twice daily reduced the mean vegetation bacterial density by 7.5 log 10 CFU/g, a result that was significantly better than that for either regimen alone ( P S. aureus aortic valve endocarditis than vancomycin alone.
Andrés J. García - One of the best experts on this subject based on the ideXlab platform.
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hydrogel delivery of Lysostaphin eliminates orthopedic implant infection by staphylococcus aureus and supports fracture healing
Proceedings of the National Academy of Sciences of the United States of America, 2018Co-Authors: Christopher T Johnson, James A Wroe, Rachit Agarwal, Karen E Martin, Rodney M Donlan, Lars F Westblade, Robert E. Guldberg, Andrés J. GarcíaAbstract:Orthopedic implant infections are a significant clinical problem, with current therapies limited to surgical debridement and systemic antibiotic regimens. Lysostaphin is a bacteriolytic enzyme with high antistaphylococcal activity. We engineered a Lysostaphin-delivering injectable PEG hydrogel to treat Staphylococcus aureus infections in bone fractures. The injectable hydrogel formulation adheres to exposed tissue and fracture surfaces, ensuring efficient, local delivery of Lysostaphin. Lysostaphin encapsulation within this synthetic hydrogel maintained enzyme stability and activity. Lysostaphin-delivering hydrogels exhibited enhanced antibiofilm activity compared with soluble Lysostaphin. Lysostaphin-delivering hydrogels eradicated S. aureus infection and outperformed prophylactic antibiotic and soluble Lysostaphin therapy in a murine model of femur fracture. Analysis of the local inflammatory response to infections treated with Lysostaphin-delivering hydrogels revealed indistinguishable differences in cytokine secretion profiles compared with uninfected fractures, demonstrating clearance of bacteria and associated inflammation. Importantly, infected fractures treated with Lysostaphin-delivering hydrogels fully healed by 5 wk with bone formation and mechanical properties equivalent to those of uninfected fractures, whereas fractures treated without the hydrogel carrier were equivalent to untreated infections. Finally, Lysostaphin-delivering hydrogels eliminate methicillin-resistant S. aureus infections, supporting this therapy as an alternative to antibiotics. These results indicate that Lysostaphin-delivering hydrogels effectively eliminate orthopedic S. aureus infections while simultaneously supporting fracture repair.
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hydrogel delivery of Lysostaphin eliminates orthopedic implant infection by staphylococcus aureus and supports fracture healing
Proceedings of the National Academy of Sciences of the United States of America, 2018Co-Authors: Christopher T Johnson, James A Wroe, Rachit Agarwal, Karen E Martin, Rodney M Donlan, Lars F Westblade, Robert E. Guldberg, Andrés J. GarcíaAbstract:Orthopedic implant infections are a significant clinical problem, with current therapies limited to surgical debridement and systemic antibiotic regimens. Lysostaphin is a bacteriolytic enzyme with high antistaphylococcal activity. We engineered a Lysostaphin-delivering injectable PEG hydrogel to treat Staphylococcus aureus infections in bone fractures. The injectable hydrogel formulation adheres to exposed tissue and fracture surfaces, ensuring efficient, local delivery of Lysostaphin. Lysostaphin encapsulation within this synthetic hydrogel maintained enzyme stability and activity. Lysostaphin-delivering hydrogels exhibited enhanced antibiofilm activity compared with soluble Lysostaphin. Lysostaphin-delivering hydrogels eradicated S. aureus infection and outperformed prophylactic antibiotic and soluble Lysostaphin therapy in a murine model of femur fracture. Analysis of the local inflammatory response to infections treated with Lysostaphin-delivering hydrogels revealed indistinguishable differences in cytokine secretion profiles compared with uninfected fractures, demonstrating clearance of bacteria and associated inflammation. Importantly, infected fractures treated with Lysostaphin-delivering hydrogels fully healed by 5 wk with bone formation and mechanical properties equivalent to those of uninfected fractures, whereas fractures treated without the hydrogel carrier were equivalent to untreated infections. Finally, Lysostaphin-delivering hydrogels eliminate methicillin-resistant S. aureus infections, supporting this therapy as an alternative to antibiotics. These results indicate that Lysostaphin-delivering hydrogels effectively eliminate orthopedic S. aureus infections while simultaneously supporting fracture repair.
Qingshan Huang - One of the best experts on this subject based on the ideXlab platform.
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Images of morphologies of MC 3T3-E1 cells on the top surface of non-Lysostaphin cement sample (a) and Lysostaphin-loaded cement sample (A) when cultured at 37°C for 3 days, and cells deeper inside the pores of non-Lysostaphin cement sample (b) and Lysostaphin-loaded cement sample (B) when cultured at 37°C for 6 days.
2014Co-Authors: Bai Xue, Jien Zhang, Cheng Zhang, Yihan Wang, Jincheng Wang, Zhizhong Cao, Qingshan HuangAbstract:Images of morphologies of MC 3T3-E1 cells on the top surface of non-Lysostaphin cement sample (a) and Lysostaphin-loaded cement sample (A) when cultured at 37°C for 3 days, and cells deeper inside the pores of non-Lysostaphin cement sample (b) and Lysostaphin-loaded cement sample (B) when cultured at 37°C for 6 days.
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MTT assay results of MC 3T3-E1 cells cultured in media incubated with normal saline, non-Lysostaphin cement sample, Lysostaphin-loaded cement sample over different times.
2014Co-Authors: Bai Xue, Jien Zhang, Cheng Zhang, Yihan Wang, Jincheng Wang, Zhizhong Cao, Qingshan HuangAbstract:Cells cultured in media alone were used as control. Non-lyso cement is non-Lysostaphin cement sample. Lyso-loaded cement is Lysostaphin-loaded cement sample.
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Fluorescence images of in vivo movement of Qdots (a), Qdot-labeled Lysostaphin (b), Qdots from cement samples (c) or Qdot-labeled Lysostaphin from cement samples (d) after subcutaneous injection in nude mice.
2014Co-Authors: Bai Xue, Jien Zhang, Cheng Zhang, Yihan Wang, Jincheng Wang, Zhizhong Cao, Qingshan HuangAbstract:Fluorescence images of in vivo movement of Qdots (a), Qdot-labeled Lysostaphin (b), Qdots from cement samples (c) or Qdot-labeled Lysostaphin from cement samples (d) after subcutaneous injection in nude mice.
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Serial setting reactions of Lysostaphin-loaded cement sample.
2014Co-Authors: Bai Xue, Jien Zhang, Cheng Zhang, Yihan Wang, Jincheng Wang, Zhizhong Cao, Qingshan HuangAbstract:Serial setting reactions of Lysostaphin-loaded cement sample.
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Cumulative and individual release profiles of Lysostaphin released from cement samples over time.
2014Co-Authors: Bai Xue, Jien Zhang, Cheng Zhang, Yihan Wang, Jincheng Wang, Zhizhong Cao, Qingshan HuangAbstract:Cumulative and individual release profiles of Lysostaphin released from cement samples over time.
Friedrich Gotz - One of the best experts on this subject based on the ideXlab platform.
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the phage lytic proteins from the staphylococcus aureus bacteriophage vb_saus phiipla88 display multiple active catalytic domains and do not trigger staphylococcal resistance
PLOS ONE, 2013Co-Authors: Lorena Rodriguezrubio, Friedrich Gotz, David M Donovan, Beatriz Martinez, Ana Rodriguez, Pilar GarciaAbstract:The increase in antibiotic resistance world-wide revitalized the interest in the use of phage lysins to combat pathogenic bacteria. In this work, we analyzed the specific cleavage sites on the staphylococcal peptidoglycan produced by three phage lytic proteins. The investigated cell wall lytic enzymes were the endolysin LysH5 derived from the S. aureus bacteriophage vB_SauS-phi-IPLA88 (phi-IPLA88) and two fusion proteins between Lysostaphin and the virion-associated peptidoglycan hydrolase HydH5 (HydH5SH3b and HydH5Lyso). We determined that all catalytic domains present in these proteins were active. Additionally, we tested for the emergence of resistant Staphylococcus aureus to any of the three phage lytic proteins constructs. Resistant S. aureus could not be identified after 10 cycles of bacterial exposure to phage lytic proteins either in liquid or plate cultures. However, a quick increase in Lysostaphin resistance (up to 1000-fold in liquid culture) was observed. The lack of resistant development supports the use of phage lytic proteins as future therapeutics to treat staphylococcal infections.
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Modular organization and resistance development of lytic enzymes.
2013Co-Authors: Lorena Rodríguez-rubio, Friedrich Gotz, David M Donovan, Beatriz Martinez, Ana Rodriguez, Pilar GarciaAbstract:A) Modular organization of lytic enzymes containing one (Lysostaphin), two (LysH5 and HydH5SH3b) or three (HydH5Lyso) catalytic domains. Large black box: endopeptidase domain; Diagonal stripes: SH3b domain; Grey box: CHAP domain; Black dots: amidase-2 domain; Horizontal stripes: LYZ2 domain. Homology: 54% between LysH5 SH3b and Lysostaphin SH3b domains; 30% between LysH5 CHAP and HydH5 CHAP domains. B) Resistance development in solid medium. Plate lysis method using 1∶2 serial dilutions of each protein spotted onto a S. aureus Sa9 lawn. C) Resistance development in liquid medium. Minimal Inhibitory Concentration (MIC) repeated exposure assay using 1∶2 serial dilutions of each protein added to 5×105 CFU/well of S. aureus Sa9. In both assays, cells surviving at ½ MIC were used as an inoculum for each subsequent round of exposure. The change in susceptibility is presented as fold change from day 1 to 10 of exposure. Error bars are the means ± standard deviations of two independent assays.
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influence of lif the Lysostaphin immunity factor on acceptors of surface proteins and cell wall sorting efficiency in staphylococcus carnosus
Journal of Bacteriology, 1998Co-Authors: Andreas Strauss, Gunther Thumm, Friedrich GotzAbstract:Proteins harboring a C-terminal cell wall sorting signal are covalently linked to pentaglycine acceptors within the staphylococcal peptidoglycan. This pentaglycine was modified when the Lysostaphin immunity factor (Lif) of Staphylococcus simulans was expressed in Staphylococcus carnosus, likely by the exchange of two glycine residues for serine residues. A reporter protein was efficiently linked to the modified acceptor, indicating that the sorting reaction is not strictly dependent on the wild-type structures of the acceptors.
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studies on proLysostaphin processing and characterization of the Lysostaphin immunity factor lif of staphylococcus simulans biovar staphylolyticus
Molecular Microbiology, 1997Co-Authors: Gunther Thumm, Friedrich GotzAbstract:Lysostaphin is an extracellular glycylglycine endopeptidase produced by Staphylococcus simulans biovar staphylolyticus ATCC1362 that lyses staphylococcal cells by hydrolysing the polyglycine interpeptide bridges of the peptidoglycan. Renewed analysis of the sequence of the Lysostaphin gene (lss), and the sequencing of the amino-terminus of purified proLysostaphin and of mature Lysostaphin revealed that Lysostaphin is organized as a preproprotein of 493 amino acids (aa), with a signal peptide consisting of 36 aa, a propeptide of 211 aa from which 195 aa are organized in 15 tandem repeats of 13 aa length, and a mature protein of 246 aa. ProLysostaphin is processed in the culture supernatant of S. simulans biovar staphylolyticus by an extracellular cysteine protease. Although proLysostaphin was staphylolytically active, the mature Lysostaphin was about 4.5-fold more active. The controlled expression in Staphylococcus carnosus of lss and lss with deletions in the prepropeptide region indicated that the tandem repeats of the propeptide are not necessary for protein export or activation of Lss, but keep Lss in a less active state. Intracellularly expressed pro- and mature Lysostaphin exert staphylolytic activity in cell-free extracts, but do not affect growth of the corresponding clones. We characterized a Lysostaphin immunity factor gene (lif) which is located in the opposite direction to lss. The expression of lif in S. carnosus led to an increase in the serine/glycine ratio of the interpeptide bridges of peptidoglycan from 2 to 35%, suggesting that Lysostaphin immunity depends on serine incorporation into the interpeptide bridge. If, in addition to lif, lss is co-expressed the serine/glycine ratio is further increased to 58%, suggesting that Lss selects for optimal serine incorporation. Lif shows similarity to FemA and FemB proteins, which are involved in the biosynthesis of the glycine interpeptide bridge of staphylococcal peptidoglycan. In contrast to that of Lif, the production of FemA and FemB in S. carnosus does not cause Lysostaphin immunity. The putative tRNASer gene located downstream of lss had no recognizable influence on Lysostaphin immunity. lss and lif are flanked by insertion sequences, suggesting that S. simulans biovar staphylolyticus received lif and lss by horizontal gene transfer.
