scholarly journals Protection of Bacillus pumilus Spores by Catalases

2012 ◽  
Vol 78 (18) ◽  
pp. 6413-6422 ◽  
Author(s):  
Aleksandra Checinska ◽  
Malcolm Burbank ◽  
Andrzej J. Paszczynski
Keyword(s):  
Hydrogen Peroxide ◽  
Bacillus Pumilus ◽  
Toxic Chemicals ◽  
Spore Coat ◽  
Synergistic Activity ◽  
Gaseous Oxygen ◽  
Content Type ◽  
Enhanced Resistance ◽  
Oxygen Gas ◽  
Coat Layer

ABSTRACTBacillus pumilusSAFR-032, isolated at spacecraft assembly facilities of the National Aeronautics and Space Administration Jet Propulsion Laboratory, is difficult to kill by the sterilization method of choice, which uses liquid or vapor hydrogen peroxide. We identified two manganese catalases, YjqC and BPUM_1305, in spore protein extracts of severalB. pumilusstrains by using PAGE and mass spectrometric analyses. While the BPUM_1305 catalase was present in six of theB. pumilusstrains tested, YjqC was not detected in ATCC 7061 and BG-B79. Furthermore, both catalases were localized in the spore coat layer along with laccase and superoxide dismutase. Although the initial catalase activity in ATCC 7061 spores was higher, it was less stable over time than the SAFR-032 enzyme. We propose that synergistic activity of YjqC and BPUM_1305, along with other coat oxidoreductases, contributes to the enhanced resistance ofB. pumilusspores to hydrogen peroxide. We observed that the product of the catalase reaction, gaseous oxygen, forms expanding vesicles on the spore surface, affecting the mechanical integrity of the coat layer, resulting in aggregation of the spores. The accumulation of oxygen gas and aggregations may play a crucial role in limiting further exposure ofBacillispore surfaces to hydrogen peroxide or other toxic chemicals when water is present.

scholarly journals The C-Terminal Zwitterionic Sequence of CotB1 Is Essential for Biosilicification of the Bacillus cereus Spore Coat

2015 ◽  
Vol 198 (2) ◽  
pp. 276-282 ◽  
Author(s):  
Kei Motomura ◽  
Takeshi Ikeda ◽  
Satoshi Matsuyama ◽  
Mohamed A. A. Abdelhamid ◽  
Tatsuya Tanaka ◽  
...  
Keyword(s):  
Coat Protein ◽  
Bacillus Cereus ◽  
Time Course ◽  
Acid Resistance ◽  
Homologous Sequence ◽  
Spore Coat ◽  
Content Type ◽  
Coat Layer ◽  
Close Relatives ◽  
Spore Coat Protein

ABSTRACTSilica is deposited in and around the spore coat layer ofBacillus cereus, and enhances the spore's acid resistance. Several peptides and proteins, including diatom silaffin and silacidin peptides, are involved in eukaryotic silica biomineralization (biosilicification). Homologous sequence search revealed a silacidin-like sequence in the C-terminal region of CotB1, a spore coat protein ofB. cereus. The negatively charged silacidin-like sequence is followed by a positively charged arginine-rich sequence of 14 amino acids, which is remarkably similar to the silaffins. These sequences impart a zwitterionic character to the C terminus of CotB1. Interestingly, thecotB1gene appears to form a bicistronic operon with its paralog,cotB2, the product of which, however, lacks the C-terminal zwitterionic sequence. A ΔcotB1B2mutant strain grew as fast and formed spores at the same rate as wild-type bacteria but did not show biosilicification. Complementation analysis showed that CotB1, but neither CotB2 nor C-terminally truncated mutants of CotB1, could restore the biosilicification activity in the ΔcotB1B2mutant, suggesting that the C-terminal zwitterionic sequence of CotB1 is essential for the process. We found that the kinetics of CotB1 expression, as well as its localization, correlated well with the time course of biosilicification and the location of the deposited silica. To our knowledge, this is the first report of a protein directly involved in prokaryotic biosilicification.IMPORTANCEBiosilicification is the process by which organisms incorporate soluble silicate in the form of insoluble silica. Although the mechanisms underlying eukaryotic biosilicification have been intensively investigated, prokaryotic biosilicification was not studied until recently. We previously demonstrated that biosilicification occurs inBacillus cereusand its close relatives, and that silica is deposited in and around a spore coat layer as a protective coating against acid. The present study reveals that aB. cereusspore coat protein, CotB1, which carried a C-terminal zwitterionic sequence, is essential for biosilicification. Our results provide the first insight into mechanisms required for biosilicification in prokaryotes.

scholarly journals Alteration of Proteomes in First-Generation Cultures of Bacillus pumilus Spores Exposed to Outer Space

mSystems ◽  
2019 ◽  
Vol 4 (4) ◽  
Author(s):  
Abby J. Chiang ◽  
Ganesh Babu Malli Mohan ◽  
Nitin K. Singh ◽  
Parag A. Vaishampayan ◽  
Markus Kalkum ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Bacillus Pumilus ◽  
Outer Space ◽  
Space Station ◽  
Ground Control ◽  
Bacterial Survival ◽  
Planetary Protection ◽  
Content Type ◽  
Atp Production ◽  
Enhanced Resistance

ABSTRACT Bacillus pumilus SAFR-032 was originally isolated from the Jet Propulsion Lab Spacecraft Assembly Facility and thoroughly characterized for its enhanced resistance to UV irradiation and oxidative stress. This unusual resistance of SAFR-032 is of particular concern in the context of planetary protection and calls for development of novel disinfection techniques to prevent extraterrestrial contamination. Previously, spores of SAFR-032 were exposed for 18 months to a variety of space conditions on board the International Space Station to investigate their resistance to Mars-like conditions and space travel. Here, proteomic characterization of vegetative SAFR-032 cells from space-surviving spores is presented in comparison to a ground control. Vegetative cells of the first passage were processed and subjected to quantitative proteomics using tandem mass tags. Approximately 60% of all proteins encoded by SAFR-032 were identified, and 301 proteins were differentially expressed among the strains. We found that proteins predicted to be involved in carbohydrate transport/metabolism and energy production/conversion had lower abundance than those of the ground control. For three proteins, we showed that the expected metabolic activities were decreased, as expected with direct enzymatic assays. This was consistent with a decrease of ATP production in the space-surviving strains. The same space-surviving strains showed increased abundance of proteins related to survival, growth advantage, and stress response. Such alterations in the proteomes provide insights into possible molecular mechanisms of B. pumilus SAFR-032 to adapt to and resist extreme extraterrestrial environments. IMPORTANCE Spore-forming bacteria are well known for their resistance to harsh environments and are of concern for spreading contamination to extraterrestrial bodies during future life detection missions. Bacillus pumilus has been regularly isolated from spacecraft-associated surfaces and exhibited unusual resistance to ultraviolet light and other sterilization techniques. A better understanding of the mechanisms of microbial survival and enhanced resistance is essential for developing novel disinfection protocols for the purpose of planetary protection. While genomic analyses did not reveal the unique characteristics that explain elevated UV resistance of space-exposed B. pumilus, the proteomics study presented here provided intriguing insight on key metabolic changes. The observed proteomics aberrations reveal a complex biological phenomenon that plays a role in bacterial survival and adaptation under long-term exposure to outer space. This adaptive ability of microorganisms needs to be considered by those tasked with eliminating forward contamination.

scholarly journals Mechanism of Sporicidal Activity for the Synergistic Combination of Peracetic Acid and Hydrogen Peroxide

2015 ◽  
Vol 82 (4) ◽  
pp. 1035-1039 ◽  
Author(s):  
Mark J. Leggett ◽  
J. Spencer Schwarz ◽  
Peter A. Burke ◽  
Gerald McDonnell ◽  
Stephen P. Denyer ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Peracetic Acid ◽  
Bacterial Spores ◽  
Spore Coat ◽  
Synergistic Activity ◽  
Wild Type ◽  
Spore Resistance ◽  
Bacterial Endospores ◽  
Synergistic Combination ◽  
Reaction Vessels

ABSTRACTThere is still great interest in controlling bacterial endospores. The use of chemical disinfectants and, notably, oxidizing agents to sterilize medical devices is increasing. With this in mind, hydrogen peroxide (H2O2) and peracetic acid (PAA) have been used in combination, but until now there has been no explanation for the observed increase in sporicidal activity. This study provides information on the mechanism of synergistic interaction of PAA and H2O2against bacterial spores. We performed investigations of the efficacies of different combinations, including pretreatments with the two oxidizers, against wild-type spores and a range of spore mutants deficient in the spore coat or small acid-soluble spore proteins. The concentrations of the two biocides were also measured in the reaction vessels, enabling the assessment of any shift from H2O2to PAA formation. This study confirmed the synergistic activity of the combination of H2O2and PAA. However, we observed that the sporicidal activity of the combination is largely due to PAA and not H2O2. Furthermore, we observed that the synergistic combination was based on H2O2compromising the spore coat, which was the main spore resistance factor, likely allowing better penetration of PAA and resulting in the increased sporicidal activity.

scholarly journals Role of the Spore Coat Layers in Bacillus subtilis Spore Resistance to Hydrogen Peroxide, Artificial UV-C, UV-B, and Solar UV Radiation

2000 ◽  
Vol 66 (2) ◽  
pp. 620-626 ◽  
Author(s):  
Paul J. Riesenman ◽  
Wayne L. Nicholson
Keyword(s):  
Hydrogen Peroxide ◽  
Bacillus Subtilis ◽  
Uv Radiation ◽  
Spore Coat ◽  
Wild Type ◽  
Spore Resistance ◽  
Solar Uv ◽  
Coat Layer ◽  
Artificial Uv ◽  
Uv C

ABSTRACT Spores of Bacillus subtilis possess a thick protein coat that consists of an electron-dense outer coat layer and a lamellalike inner coat layer. The spore coat has been shown to confer resistance to lysozyme and other sporicidal substances. In this study, spore coat-defective mutants of B. subtilis (containing thegerE36 and/or cotE::cat mutation) were used to study the relative contributions of spore coat layers to spore resistance to hydrogen peroxide (H2O2) and various artificial and solar UV treatments. Spores of strains carrying mutations in gerE and/or cotE were very sensitive to lysozyme and to 5% H2O2, as were chemically decoated spores of the wild-type parental strain. Spores of all coat-defective strains were as resistant to 254-nm UV-C radiation as wild-type spores were. Spores possessing thegerE36 mutation were significantly more sensitive to artificial UV-B and solar UV radiation than wild-type spores were. In contrast, spores of strains possessing thecotE::cat mutation were significantly more resistant to all of the UV treatments used than wild-type spores were. Spores of strains carrying both the gerE36 andcotE::cat mutations behaved likegerE36 mutant spores. Our results indicate that the spore coat, particularly the inner coat layer, plays a role in spore resistance to environmentally relevant UV wavelengths.

scholarly journals Oxidative Stress Correlates with Wolbachia-Mediated Antiviral Protection in Wolbachia-Drosophila Associations

2015 ◽  
Vol 81 (9) ◽  
pp. 3001-3005 ◽  
Author(s):  
Zhee Sheen Wong ◽  
Jeremy C. Brownlie ◽  
Karyn N. Johnson
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Biocontrol Agent ◽  
Oxygen Species ◽  
Content Type ◽  
Host Insect ◽  
Fitness Traits ◽  
Endogenous Reactive Oxygen Species ◽  
Potential Biocontrol Agent ◽  
Definition Of

ABSTRACTWolbachiamediates antiviral protection in insect hosts and is being developed as a potential biocontrol agent to reduce the spread of insect-vectored viruses. Definition of the molecular mechanism that generates protection is important for understanding the tripartite interaction between host insect,Wolbachia, and virus. Elevated oxidative stress was previously reported for a mosquito line experimentally infected withWolbachia, suggesting that oxidative stress is important forWolbachia-mediated antiviral protection. However,Wolbachiaexperimentally introduced into mosquitoes impacts a range of host fitness traits, some of which are unrelated to antiviral protection. To explore whether elevated oxidative stress is associated with antiviral protection inWolbachia-infected insects, we analyzed oxidative stress of fiveWolbachia-infectedDrosophilalines. In flies infected with protectiveWolbachiastrains, hydrogen peroxide concentrations were 1.25- to 2-fold higher than those in paired fly lines cured ofWolbachiainfection. In contrast, there was no difference in the hydrogen peroxide concentrations in flies infected with nonprotectiveWolbachiastrains compared to flies cured ofWolbachiainfection. Using aDrosophilamutant that produces increased levels of hydrogen peroxide, we investigated whether flies with high levels of endogenous reactive oxygen species had altered responses to virus infection and found that flies with high levels of endogenous hydrogen peroxide were less susceptible to virus-induced mortality. Taken together, these results suggest that elevated oxidative stress correlates withWolbachia-mediated antiviral protection in naturalDrosophilahosts.

scholarly journals Semimechanistic Pharmacokinetic/Pharmacodynamic Modeling of Fosfomycin and Sulbactam Combination against Carbapenem-Resistant Acinetobacter baumannii

2021 ◽  
Vol 65 (5) ◽  
Author(s):  
Sazlyna Mohd Sazlly Lim ◽  
Aaron J. Heffernan ◽  
Jason A. Roberts ◽  
Fekade B. Sime
Keyword(s):  
Acinetobacter Baumannii ◽  
Treatment Options ◽  
Pharmacodynamic Modeling ◽  
Synergistic Activity ◽  
Bacterial Burden ◽  
Target Attainment ◽  
Content Type ◽  
Carbapenem Resistant ◽  
Time Kill

ABSTRACT Due to limited treatment options for carbapenem-resistant Acinetobacter baumannii (CR-AB) infections, antibiotic combinations are now considered potential treatments for CR-AB. This study aimed to explore the utility of fosfomycin-sulbactam combination (FOS/SUL) therapy against CR-AB isolates. Synergism of FOS/SUL against 50 clinical CR-AB isolates was screened using the checkerboard method. Thereafter, time-kill studies against two CR-AB isolates were performed. The time-kill data were described using a semimechanistic pharmacokinetic/pharmacodynamic (PK/PD) model. Monte Carlo simulations were then performed to estimate the probability of stasis, 1-log kill, and 2-log kill after 24 h of combination therapy. The FOS/SUL combination demonstrated a synergistic effect against 74% of isolates. No antagonism was observed. The MIC50 and MIC90 of FOS/SUL were decreased 4- to 8-fold, compared to the monotherapy MIC50 and MIC90. In the time-kill studies, the combination displayed bactericidal activity against both isolates and synergistic activity against one isolate at the highest clinically achievable concentrations. Our PK/PD model was able to describe the interaction between fosfomycin and sulbactam in vitro. Bacterial kill was mainly driven by sulbactam, with fosfomycin augmentation. FOS/SUL regimens that included sulbactam at 4 g every 8 h demonstrated a probability of target attainment of 1-log10 kill at 24 h of ∼69 to 76%, compared to ∼15 to 30% with monotherapy regimens at the highest doses. The reduction in the MIC values and the achievement of a moderate PTA of a 2-log10 reduction in bacterial burden demonstrated that FOS/SUL may potentially be effective against some CR-AB infections.

scholarly journals Efficacy of Antibiotic Combinations against Multidrug-Resistant Pseudomonas aeruginosa in Automated Time-Lapse Microscopy and Static Time-Kill Experiments

2020 ◽  
Vol 64 (6) ◽  
Author(s):  
Anna Olsson ◽  
Pikkei Wistrand-Yuen ◽  
Elisabet I. Nielsen ◽  
Lena E. Friberg ◽  
Linus Sandegren ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Time Lapse ◽  
Polymyxin B ◽  
Multidrug Resistant ◽  
Positive Interactions ◽  
Synergistic Activity ◽  
Content Type ◽  
Time Lapse Microscopy ◽  
Time Kill ◽  
Antibiotic Combination

ABSTRACT Antibiotic combination therapy is used for severe infections caused by multidrug-resistant (MDR) Gram-negative bacteria, yet data regarding which combinations are most effective are lacking. This study aimed to evaluate the in vitro efficacy of polymyxin B in combination with 13 other antibiotics against four clinical strains of MDR Pseudomonas aeruginosa. We evaluated the interactions of polymyxin B in combination with amikacin, aztreonam, cefepime, chloramphenicol, ciprofloxacin, fosfomycin, linezolid, meropenem, minocycline, rifampin, temocillin, thiamphenicol, or trimethoprim by automated time-lapse microscopy using predefined cutoff values indicating inhibition of growth (≤106 CFU/ml) at 24 h. Promising combinations were subsequently evaluated in static time-kill experiments. All strains were intermediate or resistant to polymyxin B, antipseudomonal β-lactams, ciprofloxacin, and amikacin. Genes encoding β-lactamases (e.g., blaPAO and blaOXA-50) and mutations associated with permeability and efflux were detected in all strains. In the time-lapse microscopy experiments, positive interactions were found with 39 of 52 antibiotic combination/bacterial strain setups. Enhanced activity was found against all four strains with polymyxin B used in combination with aztreonam, cefepime, fosfomycin, minocycline, thiamphenicol, and trimethoprim. Time-kill experiments showed additive or synergistic activity with 27 of the 39 tested polymyxin B combinations, most frequently with aztreonam, cefepime, and meropenem. Positive interactions were frequently found with the tested combinations, against strains that harbored several resistance mechanisms to the single drugs, and with antibiotics that are normally not active against P. aeruginosa. Further study is needed to explore the clinical utility of these combinations.

scholarly journals Is Once-Daily High-Dose Ceftriaxone plus Ampicillin an Alternative for Enterococcus faecalis Infective Endocarditis in Outpatient Parenteral Antibiotic Therapy Programs?

2020 ◽  
Vol 65 (1) ◽  
pp. e02099-20
Author(s):  
Laura Herrera-Hidalgo ◽  
Arístides de Alarcón ◽  
Luis Eduardo López-Cortes ◽  
Rafael Luque-Márquez ◽  
Luis Fernando López-Cortes ◽  
...  
Keyword(s):  
Single Dose ◽  
Infective Endocarditis ◽  
Antibiotic Therapy ◽  
Enterococcus Faecalis ◽  
High Dose ◽  
Synergistic Activity ◽  
Parenteral Antibiotic ◽  
Once Daily ◽  
Content Type ◽  
Outpatient Parenteral Antibiotic Therapy

ABSTRACTCeftriaxone administered as once-daily high-dose short infusion combined with ampicillin has been proposed for the treatment of Enterococcus faecalis infective endocarditis in outpatient parenteral antibiotic therapy programs (OPAT). This combination requires synergistic activity, but the attainment of ceftriaxone synergic concentration (Cs) with the regimen proposed for OPAT has not been studied. This phase II pharmacokinetic study enrolled healthy adult volunteers who underwent two sequential treatment phases. During phase A, volunteers received 2 g of ceftriaxone each 12 h during 24 h followed by a 7-day wash-out. Then the participants received phase B, which consisted of a single dose of 4 g of ceftriaxone. Throughout both phases, each volunteer underwent intensive pharmacokinetic (PK) sampling over 24 h. Ceftriaxone total and unbound concentrations were measured. Twelve participants were enrolled and completed both phases. Mean ceftriaxone total and free concentrations 24 h after the administration of 2 g each 12 h were 86.44 ± 25.90 mg/liter and 3.59 ± 1.35 mg/liter, respectively, and after the 4-g single dose were 34.60 ± 11.16 mg/liter and 1.40 ± 0.62 mg/liter, respectively. Only 3 (25%) patients in phase A maintained unbound plasma concentrations superior to the suggested Cs = 5 mg/liter during 24 h, and none (0%) in phase B. No grade 3 to 4 adverse events or laboratory abnormalities were observed. Ceftriaxone optimal exposure combined with ampicillin to achieve maximal synergistic activity against E. faecalis required for the treatment of infective endocarditis remains unknown. However, the administration of a single daily dose of 4 g of ceftriaxone implies a reduction in the time of exposure to the proposed Cs. (This study has been registered in the European Union Drug Regulating Authorities Clinical Trials [EudraCT] database under identifier 2017-003127-29.)

scholarly journals Role of SpoIVA ATPase Motifs during Clostridioides difficile Sporulation

2020 ◽  
Vol 202 (21) ◽  
Author(s):  
Hector Benito de la Puebla ◽  
David Giacalone ◽  
Alexei Cooper ◽  
Aimee Shen
Keyword(s):  
Bacillus Subtilis ◽  
Atp Hydrolysis ◽  
Spore Formation ◽  
Spore Form ◽  
Content Type ◽  
Clostridioides Difficile ◽  
Allele Specific ◽  
Coat Layer ◽  
Quality Control Mechanism

ABSTRACT The nosocomial pathogen Clostridioides difficile is a spore-forming obligate anaerobe that depends on its aerotolerant spore form to transmit infections. Functional spore formation depends on the assembly of a proteinaceous layer known as the coat around the developing spore. In C. difficile, coat assembly depends on the conserved spore protein SpoIVA and the clostridial-organism-specific spore protein SipL, which directly interact. Mutations that disrupt their interaction cause the coat to mislocalize and impair spore formation. In Bacillus subtilis, SpoIVA is an ATPase that uses ATP hydrolysis to drive its polymerization around the forespore. Loss of SpoIVA ATPase activity impairs B. subtilis SpoIVA encasement of the forespore and activates a quality control mechanism that eliminates these defective cells. Since this mechanism is lacking in C. difficile, we tested whether mutations in the C. difficile SpoIVA ATPase motifs impact functional spore formation. Disrupting C. difficile SpoIVA ATPase motifs resulted in phenotypes that were typically >104-fold less severe than the equivalent mutations in B. subtilis. Interestingly, mutation of ATPase motif residues predicted to abrogate SpoIVA binding to ATP decreased the SpoIVA-SipL interaction, whereas mutation of ATPase motif residues predicted to disrupt ATP hydrolysis but maintain ATP binding enhanced the SpoIVA-SipL interaction. When a sipL mutation known to reduce binding to SpoIVA was combined with a spoIVA mutation predicted to prevent SpoIVA binding to ATP, spore formation was severely exacerbated. Since this phenotype is allele specific, our data imply that SipL recognizes the ATP-bound form of SpoIVA and highlight the importance of this interaction for functional C. difficile spore formation. IMPORTANCE The major pathogen Clostridioides difficile depends on its spore form to transmit disease. However, the mechanism by which C. difficile assembles spores remains poorly characterized. We previously showed that binding between the spore morphogenetic proteins SpoIVA and SipL regulates assembly of the protective coat layer around the forespore. In this study, we determined that mutations in the C. difficile SpoIVA ATPase motifs result in relatively minor defects in spore formation, in contrast with Bacillus subtilis. Nevertheless, our data suggest that SipL preferentially recognizes the ATP-bound form of SpoIVA and identify a specific residue in the SipL C-terminal LysM domain that is critical for recognizing the ATP-bound form of SpoIVA. These findings advance our understanding of how SpoIVA-SipL interactions regulate C. difficile spore assembly.

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