scholarly journals Strategy for Mass Production of Lytic Staphylococcus Aureus Bacteriophage pSa-3: Contribution of Multiplicity of Infection

2020 ◽  
Author(s):  
Sang Guen Kim ◽  
Jun Kwon ◽  
Sib Sankar Giri ◽  
Saekil Yun ◽  
Hyoun Joong Kim ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Response Surface Methodology ◽  
Response Surface ◽  
Mass Production ◽  
Fold Increase ◽  
Resistant Bacteria ◽  
Lag Phase ◽  
Multiplicity Of Infection ◽  
Bacterial Inoculum ◽  
Phage Production

Abstract BackgroundAntibiotic-resistant bacteria have emerged as a serious problem; bacteriophages have, therefore, been proposed as a therapeutic alternative to antibiotics. Several authorities, such as pharmacopeia, FDA, have confirmed their safety, and some bacteriophages are commercially available worldwide. The demand for bacteriophages is expected to increase exponentially in the future; hence, there is an urgent need to mass-produce bacteriophages economically. Unlike the replication of non-lytic bacteriophages, lytic bacteriophages are replicated by lysing host bacteria, which leads to the termination of phage production; hence, strategies that can prolong the lysis of host bacteria in bacteria-bacteriophage co-cultures are required.ResultsIn the current study, we manipulated the inoculum concentrations of Staphylococcus aureus and phage pSa-3 (multiplicity of infection, MOI), and their energy sources to delay the bactericidal effect while optimizing phage production. We examined an increasing range of bacterial inoculum concentration (2 × 108 to 2 × 109 CFU/mL) to decrease the lag phase, in combination with a decreasing range of phage inoculum (from MOI 0.01 to 0.00000001) to delay the lysis of the host. Bacterial concentration of 2 × 108 CFU/mL and phage MOI of 0.0001 showed the maximum final phage production rate (1.68 × 1010 plaque forming unit (PFU)/mL). With this combination of phage-bacteria inoculum, we selected glycerol, glycine, and calcium as carbon, nitrogen, and divalent ion sources, respectively, for phage production. After optimization using response surface methodology, the final concentration of lytic Staphylococcus phage was 8.63 × 1010 ± 9.71 × 109 PFU/mL (5.13 fold increase).ConclusionsTherefore, Staphylococcus phage pSa-3 production can be maximized by increasing the bacterial inoculum and reducing the seeding phage MOI, which this combinatorial strategy could decrease phage production time. Further, we suggest that response surface methodology has the potential for optimizing mass production of lytic bacteriophages.

scholarly journals Strategy for mass production of lytic Staphylococcus aureus bacteriophage pSa-3: contribution of multiplicity of infection and response surface methodology

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Sang Guen Kim ◽  
Jun Kwon ◽  
Sib Sankar Giri ◽  
Saekil Yun ◽  
Hyoun Joong Kim ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Response Surface Methodology ◽  
Response Surface ◽  
Mass Production ◽  
Fold Increase ◽  
Resistant Bacteria ◽  
Lag Phase ◽  
Multiplicity Of Infection ◽  
Bacterial Inoculum ◽  
Phage Production

Abstract Background Antibiotic-resistant bacteria have emerged as a serious problem; bacteriophages have, therefore, been proposed as a therapeutic alternative to antibiotics. Several authorities, such as pharmacopeia, FDA, have confirmed their safety, and some bacteriophages are commercially available worldwide. The demand for bacteriophages is expected to increase exponentially in the future; hence, there is an urgent need to mass-produce bacteriophages economically. Unlike the replication of non-lytic bacteriophages, lytic bacteriophages are replicated by lysing host bacteria, which leads to the termination of phage production; hence, strategies that can prolong the lysis of host bacteria in bacteria–bacteriophage co-cultures, are required. Results In the current study, we manipulated the inoculum concentrations of Staphylococcus aureus and phage pSa-3 (multiplicity of infection, MOI), and their energy sources to delay the bactericidal effect while optimizing phage production. We examined an increasing range of bacterial inoculum concentration (2 × 108 to 2 × 109 CFU/mL) to decrease the lag phase, in combination with a decreasing range of phage inoculum (from MOI 0.01 to 0.00000001) to delay the lysis of the host. Bacterial concentration of 2 × 108 CFU/mL and phage MOI of 0.0001 showed the maximum final phage production rate (1.68 × 1010 plaque forming unit (PFU)/mL). With this combination of phage–bacteria inoculum, we selected glycerol, glycine, and calcium as carbon, nitrogen, and divalent ion sources, respectively, for phage production. After optimization using response surface methodology, the final concentration of the lytic Staphylococcus phage was 8.63 × 1010 ± 9.71 × 109 PFU/mL (5.13-fold increase). Conclusions Therefore, Staphylococcus phage pSa-3 production can be maximized by increasing the bacterial inoculum and reducing the seeding phage MOI, and this combinatorial strategy could decrease the phage production time. Further, we suggest that response surface methodology has the potential for optimizing the mass production of lytic bacteriophages.

scholarly journals Chitinase from a Novel Strain ofSerratia marcescensJPP1 for Biocontrol of Aflatoxin: Molecular Characterization and Production Optimization Using Response Surface Methodology

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Kai Wang ◽  
Pei-sheng Yan ◽  
Li-xin Cao
Keyword(s):  
Response Surface Methodology ◽  
Response Surface ◽  
Pcr Amplification ◽  
Fold Increase ◽  
Antagonistic Activity ◽  
Medium Composition ◽  
Production Optimization ◽  
Reading Frame ◽  
Mycolytic Enzymes ◽  
Burman Design

Chitinase is one of the most important mycolytic enzymes with industrial significance, and produced by a number of organisms. A chitinase producing isolateSerratia marcescensJPP1 was obtained from peanut hulls in Jiangsu Province, China, and exhibited antagonistic activity against aflatoxins. In this study, we describe the optimization of medium composition with increased production of chitinase for the selected bacteria using statistical methods: Plackett-Burman design was applied to find the key ingredients, and central composite design of response surface methodology was used to optimize the levels of key ingredients for the best yield of chitinase. Maximum chitinase production was predicted to be 23.09 U/mL for a 2.1-fold increase in medium containing 12.70 g/L colloidal chitin, 7.34 g/L glucose, 5.00 g/L peptone, 1.32 g/L (NH4)2SO4, 0.7 g/L K2HPO4, and 0.5 g/L MgSO4·7H2O. Polymerase chain reaction (PCR) amplification of the JPP1 chitinase gene was performed and obtained a 1,789 bp nucleotide sequence; its open reading frame encoded a protein of 499 amino acids named as ChiBjp.

scholarly journals Application of Response Surface Methodology for Optimizing Arginine Deiminase Production Medium forEnterococcus faeciumsp. GR7

2013 ◽  
Vol 2013 ◽  
pp. 1-12 ◽  
Author(s):  
Baljinder Kaur ◽  
Rajinder Kaur
Keyword(s):  
Response Surface Methodology ◽  
Central Composite Design ◽  
Response Surface ◽  
Specific Activity ◽  
Basal Medium ◽  
Fold Increase ◽  
Arginine Deiminase ◽  
Arginine Metabolism ◽  
Central Composite ◽  
Fermentation Media

Arginine metabolism inEnterococcus faeciumsp. GR7 was enhanced via arginine deiminase pathway. Process parameters including fermentation media and environmental conditions were optimized using independent experiments and response surface methodology (central composite design). Fermentation media (EAPM) were optimized using independent experiments which resulted in 4-fold increase in arginine deiminase specific activity as compared to basal medium. To further enhance arginine deiminase activity inE. faeciumsp. GR7 and biomass production including a five-level central composite design (CCD) was employed to study the interactive effect of three-process variables. Response surface methodology suggested a quadratic model which was further validated experimentally where it showed approximately 15-fold increase in arginine metabolism (in terms of arginine deiminase specific activity) over basal medium. By solving the regression equation and analyzing the response surface cartons, optimal concentrations of the media components (g/L) were determined as arginine 20.0; tryptone 15.0; lactose 10.0; K2HPO43.0; NaCl 1.0, MnSO40.6 mM; Tween 80 1%; pH 6.0 for achieving specific arginine deiminase activity of 4.6 IU/mG with concomitant biomass production of 12.1 mg/L. The model is significant as the coefficient of determination (R2) was 0.87 to 0.90 for all responses. Enhanced arginine deiminase yield fromE. faecium, a GRAS lactic acid bacterial strain, is desirable to explorein vitrotherapeutic potential of the arginine metabolizingE. faeciumsp. GR7.

scholarly journals Application of the response surface methodology for the evaluation of Staphylococcus aureus inhibition with Ag/TiO2 nanoparticles

2021 ◽  
Author(s):  
M.A. Olivares-Ramírez ◽  
Leticia López-Zamora ◽  
M.J. Peña-Juárez ◽  
E.J. Gutiérrez-Castañeda ◽  
J.A. Gonzalez-Calderon
Keyword(s):  
Staphylococcus Aureus ◽  
Response Surface Methodology ◽  
Response Surface ◽  
Statistical Significance ◽  
Antimicrobial Effect ◽  
Quadratic Model ◽  
Coefficient Of Determination ◽  
Poly Lactic Acid ◽  
P Value ◽  
Zone Of Inhibition

Abstract The present work shows the implementation of the Response Surface Methodology (RSM), fed by an experimental Central Composite Design (CCD) to find the conditions that allow maximizing the inhibition of the microorganism Staphylococcus aureus with nanoparticles of TiO2 silanized with 3-Aminopropyltriethoxysilane (APTES) and doped with Ag. In addition, Poly(lactic) acid composites were prepared with these Ag/TiO2 nanoparticles with the aim to confer their antimicrobial effect. The independent variables considered were pH, AgNO3/TiO2 ratio (% w/w), and TiO2 nanoparticles concentration (g/250 mL), and as the variable of response, the length of the diameter of the halo or zone of inhibition presented by the microorganism (mm). Statistical analysis found that maximization of S. aureus inhibition occurs at intermediate levels with a value of 10 for pH and 5 g of TiO2 solids, while for the concentration of AgNO3 high levels are required, greater than 10% w/w. Likewise, the statistical significance was determined using the Student's t-test and the p-value; it was found that the significant effect corresponds to the concentration of AgNO3, so a second experimental CCD design equirradial with two factors was considered, estimating AgNO3 concentration and TiO2 amount, the pH at constant 10 value. The second experimental design indicated that maximization in S. aureus inhibition occurs at an AgNO3 concentration between 20-25% w/w with high amounts of TiO2 solids (7-8 g), with a resulting zone of inhibition between 26-28 mm. The quadratic model obtained, which represents the relationship between the length of the zone of inhibition with the variables considered, shows an adjustment of experimental data with a coefficient of determination (R2) of 0.82.

scholarly journals Statistical optimization of culture medium composition for enhanced zeaxanthin production by Cyanophycean microalgae Trichodesmium thiebautii (NIOT 152)

2020 ◽  
Vol 13 (41) ◽  
pp. 4307-4318
Author(s):  
Priyanka Srinivasan ◽  
Keyword(s):  
Response Surface Methodology ◽  
Response Surface ◽  
Sea Water ◽  
Fold Increase ◽  
Medium Composition ◽  
Statistical Optimization ◽  
Age Related Macular Degeneration ◽  
Medium Components ◽  
Age Related ◽  
Trichodesmium Thiebautii

Background/Objectives: Zeaxanthin is a xanthophyll carotenoid revered for its role in the prevention of age related macular degeneration. The study evaluated the zeaxanthin accumulation of the marine Cyanophycean alga Trichodesmium thiebautii (NIOT 152). A sequential statistical technique was applied to optimize the Artificial Sea Water nutrient medium (ASN-III) components for enhancing the zeaxanthin accumulation in T. thiebautii. Methods: A two-level statistical approach involving Plackett-Burman (PB) design to screen the most important nutrients influencing the zeaxanthin accumulation followed by Response surface methodology (RSM) was employed. The results of PB design revealed sodium nitrate, disodium EDTA, magnesium sulphate and sodium carbonate as the crucial medium components for increasing zeaxanthin accumulation. Further, RSM was employed to study the interaction between these factors and identified an optimum concentration of the ingredients for higher zeaxanthin production. Findings: The optimized medium components resulted in 2.33 fold increase in zeaxanthin accumulation (4.3 ± 1.29 mg L-1) as compared to ASN III medium (1.84 ± 0.12 mg L-1). Novelty: There are only few studies on laboratory cultured Trichodesmium and only very few reports are available regarding pigment production from Trichodesmium sp. The present study successfully demonstrated the statistical optimization of ASN III medium to improve zeaxanthin accumulation by Trichodesmium thiebautii. Keywords: ASN III medium; zeaxanthin; Trichodesmium thiebautii; Plackett-Burman; response surface methodology REFERENCE

scholarly journals Application of Response Surface Methodology to Evaluate Photodynamic Inactivation Mediated by Eosin Y and 530 nm LED against Staphylococcus aureus

Antibiotics ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 125 ◽  
Author(s):  
Adriele R. Santos ◽  
Alex F. da Silva ◽  
Andréia F. P. Batista ◽  
Camila F. Freitas ◽  
Evandro Bona ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Response Surface Methodology ◽  
Response Surface ◽  
Foodborne Pathogens ◽  
Irradiation Time ◽  
Eosin Y ◽  
Efficient Tool ◽  
Order Polynomial ◽  
Negative Effect ◽  
The Relationship

Photodynamic antimicrobial chemotherapy (PAC) is an efficient tool for inactivating microorganisms. This technique is a good approach to inactivate the foodborne microorganisms, which are responsible for one of the major public health concerns worldwide—the foodborne diseases. In this work, response surface methodology (RSM) was used to evaluate the interaction of Eosin Y (EOS) concentration and irradiation time on Staphylococcus aureus counts and a sequence of designed experiments to model the combined effect of each factor on the response. A second-order polynomial empirical model was developed to describe the relationship between EOS concentration and irradiation time. The results showed that the derived model could predict the combined influences of these factors on S. aureus counts. The agreement between predictions and experimental observations (R2adj = 0.9159, p = 0.000034) was also observed. The significant terms in the model were the linear negative effect of photosensitizer (PS) concentration, followed by the linear negative effect of irradiation time, and the quadratic negative effect of PS concentration. The highest reductions in S. aureus counts were observed when applying a light dose of 9.98 J/cm2 (498 nM of EOS and 10 min. irradiation). The ability of the evaluated model to predict the photoinactivation of S. aureus was successfully validated. Therefore, the use of RSM combined with PAC is a promising approach to inactivate foodborne pathogens.

Optimization of fermentation parameters in phage production using response surface methodology

2012 ◽  
Vol 39 (10) ◽  
pp. 1515-1522 ◽  
Author(s):  
Sung-Hye H. Grieco ◽  
Ann Y. K. Wong ◽  
W. Scott Dunbar ◽  
Ross T. A. MacGillivray ◽  
Susan B. Curtis
Keyword(s):  
Response Surface Methodology ◽  
Response Surface ◽  
Phage Production ◽  
Fermentation Parameters ◽  
Optimization Of Fermentation

scholarly journals Investigation of the Potential for Mutational Resistance to XF-73, Retapamulin, Mupirocin, Fusidic Acid, Daptomycin, and Vancomycin in Methicillin-ResistantStaphylococcus aureusIsolates during a 55-Passage Study

2010 ◽  
Vol 55 (3) ◽  
pp. 1177-1181 ◽  
Author(s):  
David J. Farrell ◽  
Marion Robbins ◽  
William Rhys-Williams ◽  
William G. Love
Keyword(s):  
Staphylococcus Aureus ◽  
Fusidic Acid ◽  
Bacterial Resistance ◽  
Point Mutations ◽  
Hospital Setting ◽  
Fold Increase ◽  
Cross Resistance ◽  
Resistant Bacteria ◽  
Sequencing Analysis ◽  
Antibacterial Drugs

ABSTRACTXF-73 is a dicationic porphyrin drug with rapid Gram-positive antibacterial activity currently undergoing clinical trials for the nasal decolonization ofStaphylococcus aureus, including methicillin-resistantStaphylococcus aureus(MRSA). In multistep (55-passage) resistance selection studies in the presence of subinhibitory concentrations of XF-73, retapamulin, mupirocin, fusidic acid, and vancomycin against four Network on Antimicrobial Resistance inStaphylococcus aureusMRSA strains, there was no >4-fold increase in the MIC for XF-73 after 55 passages. In contrast, there was an increase in the MICs for retapamulin (from 0.25 μg/ml to 4 to 8 μg/ml), for mupirocin (from 0.12 μg/ml to 16 to 512 μg/ml), for fusidic acid (from 0.12 μg/ml to 256 μg/ml), and for vancomycin (from 1 μg/ml to 8 μg/ml in two of the four strains tested). Further investigations usingS. aureusNRS384 (USA300) and daptomycin demonstrated a 64-fold increase in the MIC after 55 passages (from 0.5 μg/ml to 32 μg/ml) with a >4-fold increase in the MIC obtained after only five passages. Sequencing analysis of selected isolates confirmed previously reported point mutations associated with daptomycin resistance. No cross-resistance to XF-73 was observed with the daptomycin-resistant strains, suggesting that whereas the two drugs act on the bacterial cell membrane, their specific site of action differs. XF-73 thus represents the first in a new class of antibacterial drugs, which (unlike the comparator antibiotics) after 55 passages exhibited a ≤4-fold increase in MIC against the strains tested. Antibacterial drugs with a low propensity for inducing bacterial resistance are much needed for the prevention and treatment of multidrug-resistant bacteria both within and outside the hospital setting.

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