Comparison of Predictive Models for Growth of Parent and Green Fluorescent Protein–Producing Strains of Salmonella†

2003 ◽  
Vol 66 (2) ◽  
pp. 200-207 ◽  
Author(s):  
T. P. OSCAR
Keyword(s):  
Growth Rate ◽  
Growth Kinetics ◽  
Predictive Models ◽  
Fluorescent Protein ◽  
Specific Growth ◽  
Growth Parameters ◽  
Contaminated Food ◽  
Green Fluorescent ◽  
Kinetics Of ◽  
Maximum Population Density

The green fluorescent protein (GFP) from the jellyfish Aequorea victoria can be expressed in, and used to follow the fate of, Salmonella in microbiologically complex ecosystems such as food. As a first step in the evaluation of GFP as a tool for the development of predictive models for naturally contaminated food, the present study was undertaken to compare the growth kinetics of parent and GFP-producing strains of Salmonella. A previously established sterile chicken burger model system was used to compare the growth kinetics of stationary-phase cells of parent and GFP strains of Salmonella Enteritidis, Salmonella Typhimurium, and Salmonella Dublin. Growth curves for constant temperatures from 10 to 48°C were fit to a two- or three-phase linear model to determine lag time, specific growth rate, and maximum population density. Secondary models for the growth parameters as a function of temperature were generated and compared between the parent and GFP strain pairs. The effects of GFP on the three growth parameters were significant and were affected by serotype and incubation temperature. The expression of GFP reduced specific growth rate and maximum population density while having only a small effect on the lag times of the three serotypes. The results of this study indicate that the growth kinetics of the GFP strains tested were different from those of the parent strains and thus would not be good marker strains for the development of predictive models for naturally contaminated food.

Growth Modeling of Listeria monocytogenes in Pasteurized Liquid Egg

2013 ◽  
Vol 76 (9) ◽  
pp. 1549-1556 ◽  
Author(s):  
MIHO OHKOCHI ◽  
SHIGENOBU KOSEKI ◽  
MASAAKI KUNOU ◽  
KATSUAKI SUGIURA ◽  
HIROKAZU TSUBONE
Keyword(s):  
Listeria Monocytogenes ◽  
Specific Growth ◽  
Storage Temperature ◽  
Growth Parameters ◽  
Root Model ◽  
Natural Flora ◽  
Mean Square Errors ◽  
Kinetics Of ◽  
Storage Temperatures ◽  
Maximum Population Density

The growth kinetics of Listeria monocytogenes and natural flora in commercially produced pasteurized liquid egg was examined at 4.1 to 19.4°C, and a growth simulation model that can estimate the range of the number of L. monocytogenes bacteria was developed. The experimental kinetic data were fitted to the Baranyi model, and growth parameters, such as maximum specific growth rate (μmax), maximum population density (Nmax), and lag time (λ), were estimated. As a result of estimating these parameters, we found that L. monocytogenes can grow without spoilage below 12.2°C, and we then focused on storage temperatures below 12.2°C in developing our secondary models. The temperature dependency of the μmax was described by Ratkowsky's square root model. The Nmax of L. monocytogenes was modeled as a function of temperature, because the Nmax of L. monocytogenes decreased as storage temperature increased. A tertiary model of L. monocytogenes was developed using the Baranyi model and μmax and Nmax secondary models. The ranges of the numbers of L. monocytogenes bacteria were simulated using Monte Carlo simulations with an assumption that these parameters have variations that follow a normal distribution. Predictive simulations under both constant and fluctuating temperature conditions demonstrated a high accuracy, represented by root mean square errors of 0.44 and 0.34, respectively. The predicted ranges also seemed to show a reasonably good estimation, with 55.8 and 51.5% of observed values falling into the prediction range of the 25th to 75th percentile, respectively. These results suggest that the model developed here can be used to estimate the kinetics and range of L. monocytogenes growth in pasteurized liquid egg under refrigerated temperature.

Transformation of Escherichia coli K-12 with a High-Copy Plasmid Encoding the Green Fluorescent Protein Reduces Growth: Implications for Predictive Microbiology†

2006 ◽  
Vol 69 (2) ◽  
pp. 276-281 ◽  
Author(s):  
T. P. OSCAR ◽  
K. DULAL ◽  
D. BOUCAUD
Keyword(s):  
Escherichia Coli ◽  
Green Fluorescent Protein ◽  
Predictive Models ◽  
Fluorescent Protein ◽  
Inducible Promoter ◽  
E Coli ◽  
Contaminated Food ◽  
Green Fluorescent ◽  
K 12 ◽  
High Copy Plasmid

The green fluorescent protein (GFP) of the jellyfish Aequorea victoria has been widely used as a biomarker and has potential for use in developing predictive models for growth of pathogens on naturally contaminated food. However, constitutive production of GFP can reduce growth of transformed strains. Consequently, a high-copy plasmid with gfp under the control of a tetracycline-inducible promoter (pTGP) was constructed. The plasmid was first introduced into a tetracycline-resistant strain of Escherichia coli K-12 to propagate it for subsequent transformation of tetracycline-resistant strains of Salmonella. In contrast to transformed E. coli K-12, which only fluoresced in response to tetracycline, transformed Salmonella fluoresced maximally without tetracycline induction of gfp. Although pTGP did not function as intended in Salmonella, growth of parent and GFP E. coli K-12 was compared to test the hypothesis that induction of GFP production reduced growth. Although GFP production was not induced during growth on sterile chicken in the absence of tetracycline, maximum specific growth rate (μmax) of GFP E. coli K-12 was reduced 40 to 50% (P < 0.05) at 10, 25, and 40°C compared with the parent strain. When growth of parent and GFP strains of E. coli K-12 was compared in sterile broth at 40°C, μmax and maximum population density of the GFP strain were reduced (P < 0.05) to the same extent (50 to 60%) in the absence and presence of tetracycline. These results indicated that transformation reduced growth of E. coli K-12 independent of gfp induction. Thus, use of a low-copy plasmid or insertion of gfp into the chromosome may be required to construct valid strains for development of predictive models for growth of pathogens on naturally contaminated food.

scholarly journals Growth Kinetics of Hyphomicrobium and Thiobacillus spp. in Mixed Cultures Degrading Dimethyl Sulfide and Methanol

2010 ◽  
Vol 76 (16) ◽  
pp. 5423-5431 ◽  
Author(s):  
Alexander C. Hayes ◽  
Steven N. Liss ◽  
D. Grant Allen
Keyword(s):  
Growth Rate ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Specific Growth Rate ◽  
Growth Kinetics ◽  
Growth Rates ◽  
Dimethyl Sulfide ◽  
Specific Growth ◽  
Rrna Gene ◽  
Kinetics Of

ABSTRACT The growth kinetics of Hyphomicrobium spp. and Thiobacillus spp. on dimethyl sulfide (DMS) and methanol (in the case of Hyphomicrobium spp.) in an enrichment culture created from a biofilter cotreating DMS and methanol were studied. Specific growth rates of 0.099 h−1 and 0.11 h−1 were determined for Hyphomicrobium spp. and Thiobacillus spp., respectively, growing on DMS at pH 7. These specific growth rates are double the highest maximum specific growth rate for bacterial growth on DMS reported to date in the literature. When the pH of the medium was decreased from pH 7 to pH 5, the specific growth rate of Hyphomicrobium spp. decreased by 85%, with a near 100-fold decline in the yield of Hyphomicrobium 16S rRNA gene copies in the mixed culture. Through the same pH shift, the specific growth rate and 16S rRNA gene yield of Thiobacillus spp. remained similar. When methanol was used as a substrate, the specific growth rate of Hyphomicrobium spp. declined much less over the same pH range (up to 30%) while the yield of 16S rRNA gene copies declined by only 50%. Switching from an NH4 +-N-based source to a NO3 −-N-based source resulted in the same trends for the specific growth rate of these microorganisms with respect to pH. This suggests that pH has far more impact on the growth kinetics of these microorganisms than the nitrogen source. The results of these mixed-culture batch experiments indicate that the increased DMS removal rates observed in previous studies of biofilters cotreating DMS and methanol are due to the proliferation of DMS-degrading Hyphomicrobium spp. on methanol at pH levels not conducive to high growth rates on DMS alone.

Growth kinetics of Escherichia coli with galactose and several other sugars in carbon-limited chemostat culture

1999 ◽  
Vol 46 (1) ◽  
pp. 72-80 ◽  
Author(s):  
Urs Lendenmann ◽  
Mario Snozzi ◽  
Thomas Egli
Keyword(s):  
Escherichia Coli ◽  
Growth Rate ◽  
Steady State ◽  
Specific Growth Rate ◽  
Growth Kinetics ◽  
Specific Growth ◽  
Chemostat Culture ◽  
Monod Model ◽  
E Coli ◽  
Kinetics Of

Kinetic models for microbial growth describe the specific growth rate (μ) as a function of the concentration of the growth-limiting nutrient (s) and a set of parameters. A typical example is the model proposed by Monod, where μ is related to s using substrate affinity (Ks) and the maximum specific growth rate (μmax). The preferred method to determine such parameters is to grow microorganisms in continuous culture and to measure the concentration of the growth-limiting substrate as a function of the dilution rate. However, owing to the lack of analytical methods to quantify sugars in the microgram per litre range, it has not been possible to investigate the growth kinetics of Escherichia coli in chemostat culture. Using an HPLC method able to determine steady-state concentrations of reducing sugars, we previously have shown that the Monod model adequately describes glucose-limited growth of E. coli ML30. This has not been confirmed for any other sugar. Therefore, we carried out a similar study with galactose and found steady-state concentrations between 18 and 840 μg·L-1 for dilution rates between 0.2 and 0.8·h-1, respectively. With these data the parameters of several models giving the specific growth rate as a function of the substrate concentration were estimated by nonlinear parameter estimation, and subsequently, the models were evaluated statistically. From all equations tested, the Monod model described the data best. The parameters for galactose utilisation were μmax = 0.75·h-1 and Ks = 67 μg·L-1. The results indicated that accurate Ks values can be estimated from a limited set of steady-state data when employing μmax measured during balanced growth in batch culture. This simplified procedure was applied for maltose, ribose, and fructose. For growth of E. coli with these sugars, μmax and Ks were for maltose 0.87·h-1, 100 μg·L-1; for ribose 0.57·h-1, 132 μg·L-1, and for fructose 0.70·h-1, 125 μg·L-1. Key words: monod model, continuous culture, galactose, glucose, fructose, maltose, ribose.

scholarly journals Effect of Environmental Factors on Growth Kinetics of Biocontrol Agent Bacillus thuringiensis Bacterium using 2L and 5L A+ Sartorius Stedim Biostat® Fermentation Systems

2020 ◽  
Vol 2 (6) ◽  
Author(s):  
C. S. Richardson ◽  
D. Upadhyay ◽  
S. Mandjiny ◽  
L. Holmes
Keyword(s):  
Growth Rate ◽  
Bacillus Thuringiensis ◽  
Growth Kinetics ◽  
Growth Rates ◽  
Environmental Conditions ◽  
Doubling Time ◽  
Specific Growth ◽  
Optimal Conditions ◽  
Specific Growth Rates ◽  
Kinetics Of

Bacillus thuringiensis (Bt) is a soil-dwelling, Gram-positive bacterium that is used as a biological pesticide and used to genetically engineer plants due to the toxic proteins it produces. B. thuringiensis was studied in batch cultures to determine the specific growth rates and doubling times. The purpose of this experiment was to research the growth kinetics of Bacillus thuringiensis in a 2L bioreactor and a 5L bioreactor containing growth media at different environmental conditions. Fermentation parameters were controlled by utilizing a Sartorius Stedim Biostat® A+ bioreactor system for bacterial growth. The environmental conditions included temperature, agitation, and aeration. The specific growth rates of B. thuringiensis were determined. The optimal conditions for the 2L bioreactor were 200 RPM, 30°C, 1.5 VVM, and with the highest specific growth rate 0.30 hr and the shortest doubling time 2.3 hr. For the 5L bioreactor, the optimal conditions were 150 RPM, 30°C, 1.5 VVM, and with the highest specific growth rate 1.2 hr and the fastest doubling time 0.6 hr.

scholarly journals Growth and endoglucanase activity of Acetivibrio cellulolyticus grown in three different cellulosic substrates

1999 ◽  
Vol 30 (4) ◽  
pp. 310-314 ◽  
Author(s):  
Cássia Regina Sanchez ◽  
Clarita Schvartz Peres ◽  
Heloiza Ramos Barbosa
Keyword(s):  
Growth Rate ◽  
Enzyme Activity ◽  
Growth Kinetics ◽  
Specific Growth ◽  
Carbon Sources ◽  
The Other ◽  
Crystalline Cellulose ◽  
Amorphous Cellulose ◽  
Endoglucanase Activity ◽  
Kinetics Of

The growth kinetics of Acetivibrio cellulolyticus grown in medium containing different carbon sources (cellobiose, amorphous or crystalline cellulose) was investigated. The specific growth rate was higher in cellobiose fed cultures than in the presence of the other two substrates. Endoglucanase production was greater in cultures grown on amorphous cellulose; enzyme activity increased during the stationary phase in cultures grown on crystalline cellulose.

scholarly journals Development of a Fluorescent Tool for Studying Legionella bozemanae Intracellular Infection

2021 ◽  
Vol 9 (2) ◽  
pp. 379
Author(s):  
Breanne M. Head ◽  
Christopher I. Graham ◽  
Teassa MacMartin ◽  
Yoav Keynan ◽  
Ann Karen C. Brassinga
Keyword(s):  
Growth Kinetics ◽  
Legionella Pneumophila ◽  
Fluorescent Protein ◽  
Disease Incidence ◽  
Acanthamoeba Castellanii ◽  
Infection Model ◽  
Intracellular Infection ◽  
Green Fluorescent

Legionnaires’ disease incidence is on the rise, with the majority of cases attributed to the intracellular pathogen, Legionella pneumophila. Nominally a parasite of protozoa, L. pneumophila can also infect alveolar macrophages when bacteria-laden aerosols enter the lungs of immunocompromised individuals. L. pneumophila pathogenesis has been well characterized; however, little is known about the >25 different Legionella spp. that can cause disease in humans. Here, we report for the first time a study demonstrating the intracellular infection of an L. bozemanae clinical isolate using approaches previously established for L. pneumophila investigations. Specifically, we report on the modification and use of a green fluorescent protein (GFP)-expressing plasmid as a tool to monitor the L. bozemanae presence in the Acanthamoeba castellanii protozoan infection model. As comparative controls, L. pneumophila strains were also transformed with the GFP-expressing plasmid. In vitro and in vivo growth kinetics of the Legionella parental and GFP-expressing strains were conducted followed by confocal microscopy. Results suggest that the metabolic burden imposed by GFP expression did not impact cell viability, as growth kinetics were similar between the GFP-expressing Legionella spp. and their parental strains. This study demonstrates that the use of a GFP-expressing plasmid can serve as a viable approach for investigating Legionella non-pneumophila spp. in real time.

scholarly journals AKTIVITAS INULINASE OLEH Pichia manshurica DAN FUSAN F4 PADA FERMENTASI BATCH DENGAN UMBI DAHLIA (Dahlia sp) SEBAGAI SUBSTRAT

REAKTOR ◽  
2015 ◽  
Vol 14 (3) ◽  
pp. 187 ◽  
Author(s):  
Wijanarka Wijanarka ◽  
Endang Sutariningsih Soetarto ◽  
Kumala Dewi ◽  
Ari Indrianto
Keyword(s):  
Growth Rate ◽  
Specific Growth Rate ◽  
Specific Growth ◽  
Lag Phase ◽  
Growth Media ◽  
A Value ◽  
Inulinase Activity ◽  
Pichia Manshurica ◽  
Microbial Strains ◽  
Kinetics Of

ACTIVITY OF INULINASE OF Pichia Manshuria AND FUSAN F4 ON BATCH FERMENTATION UDING DAHLIA TUBER (Dahlia sp) AS A SUBSTRATE. A dahlia tuber is one of the common inulin rich crops. Inulin is formed by units of fructans, which are polymers of D-fructose. Inulinases (EC 3.2.1.7) catalyze the hydrolysis of inulin, producing fructooligosaccharides (FOS), inulooligosaccharides (IOS), pulullan, acetone, butanol and sorbitol, therefore dahlia tubers are used as growth media. The inulin hydrolyzing activity has been reported from various microbial strains Pichia manshurica and Fusan F4 which is the result of fusion protoplast. The objective of this study was to determine the activity of inulinase Pichia manshurica and Fusan F4 on the substrate dahlia tubers. Fusan F4 to increase inulinase activity compared with Pichia manshurica and to investigate the kinetics of specific growth rate (μ) and time double (g) from of Pichia manshurica and Fusan F4. The results showed that the exponential phase occurs at 0-12 hour without a lag phase. P. manshurica has a specific growth rate (μ) of 0.18/hour with time double (g) 3.90 hours and the inulinase enzyme activity of 0.56 IU, while for Fusan F4 consecutive has a value μ of 0.20/hour, g of 3.49 hours and the activity of 0.69 IU. The conclusion of this research is to improve Fusan F4 inulinase activity and the ability has to be better than the Pichia manshurica.Umbi dahlia merupakan salah satu umbi yang mengandung inulin. Inulin merupakan polimer fruktan yang dapat dipecah oleh enzim inulinase (E.C. 3.2.1.7) menjadi fruktosa. Fruktosa merupakan bahan baku dasar untuk pembuatan FOS, IOS, pulullan, aseton dan sorbitol, oleh karena itu umbi dahlia digunakan sebagai media pertumbuhan. Enzim inulinase ini secara indigenous dimiliki oleh Pichia manshurica dan Fusan F4 yang merupakan hasil fusi protoplas.Tujuan  penelitian ini adalah  untuk mengetahui aktivitas inulinase Pichia manshurica dan Fusan F4 pada substrat umbi dahlia, Fusan F4 mampu meningkatkan aktivitas inulinase dibandingkan dengan Pichia manshurica serta untuk mengetahui kinetika kecepatan pertumbuhan specifik (µ) dan waktu generasi (g) Pichia manshurica dan Fusan F4. Hasil penelitian menunjukkan bahwa fase  eksponensial terjadi pada jam ke-0 sampai jam ke-12 tanpa diikuti fase lag, Pichia manshurica mempunyai kecepatan pertumbuhan specific (µ)  sebesar 0,18/jam dengan waktu generasi (g) 3,90 jam dan aktivitas enzim inulinase yang dihasilkan sebesar 0,56 IU, sedangkan untuk fusan F4 secara berturut-turut mempunyai nilai µ sebesar 0,20/jam, g sebesar 3,49 jam dan aktivitas sebesar 0,69 IU. Kesimpulan dari penelitian ini adalah Fusan F4 mampu meningkatkan aktivitas inulinase dan mempunyai kemampuan lebih baik dibanding dengan Pichia manshurica.

scholarly journals The sustainability of interactions between the orexin-1 receptor and β-arrestin-2 is defined by a single C-terminal cluster of hydroxy amino acids and modulates the kinetics of ERK MAPK regulation

2005 ◽  
Vol 387 (3) ◽  
pp. 573-584 ◽  
Author(s):  
Sandra MILASTA ◽  
Nicholas A. EVANS ◽  
Laura ORMISTON ◽  
Shelagh WILSON ◽  
Robert J. LEFKOWITZ ◽  
...  
Keyword(s):  
Amino Acids ◽  
Fluorescent Protein ◽  
Weak Interactions ◽  
Dependent Manner ◽  
Wild Type ◽  
Erk Mapk ◽  
Hydroxy Amino ◽  
C Terminus ◽  
Green Fluorescent ◽  
Kinetics Of

The orexin-1 receptor interacts with β-arrestin-2 in an agonist-dependent manner. In HEK-293T cells, these two proteins became co-internalized into acidic endosomes. Truncations from the C-terminal tail did not prevent agonist-induced internalization of the orexin-1 receptor or alter the pathway of internalization, although such mutants failed to interact with β-arrestin-2 in a sustained manner or produce its co-internalization. Mutation of a cluster of three threonine and one serine residue at the extreme C-terminus of the receptor greatly reduced interaction and abolished co-internalization of β-arrestin-2–GFP (green fluorescent protein). Despite the weak interactions of this C-terminally mutated form of the receptor with β-arrestin-2, studies in wild-type and β-arrestin-deficient mouse embryo fibroblasts confirmed that agonist-induced internalization of this mutant required expression of a β-arrestin. Although without effect on agonist-mediated elevation of intracellular Ca2+ levels, the C-terminally mutated form of the orexin-1 receptor was unable to sustain phosphorylation of the MAPKs (mitogen-activated protein kinases) ERK1 and ERK2 (extracellular-signal-regulated kinases 1 and 2) to the same extent as the wild-type receptor. These studies indicate that a single cluster of hydroxy amino acids within the C-terminal seven amino acids of the orexin-1 receptor determine the sustainability of interaction with β-arrestin-2, and indicate an important role of β-arrestin scaffolding in defining the kinetics of orexin-1 receptor-mediated ERK MAPK activation.

scholarly journals Hsp104 Overexpression Cures Saccharomyces cerevisiae [ PSI + ] by Causing Dissolution of the Prion Seeds

2014 ◽  
Vol 13 (5) ◽  
pp. 635-647 ◽  
Author(s):  
Yang-Nim Park ◽  
Xiaohong Zhao ◽  
Yang-In Yim ◽  
Horia Todor ◽  
Robyn Ellerbrock ◽  
...  
Keyword(s):  
Molecular Chaperone ◽  
Fluorescent Protein ◽  
Yeast Cells ◽  
Yeast Prion ◽  
Yeast Prions ◽  
Daughter Cells ◽  
Amyloid Aggregates ◽  
Mother And Daughter ◽  
Green Fluorescent ◽  
Kinetics Of

ABSTRACT The [ PSI + ] yeast prion is formed when Sup35 misfolds into amyloid aggregates. [ PSI + ], like other yeast prions, is dependent on the molecular chaperone Hsp104, which severs the prion seeds so that they pass on as the yeast cells divide. Surprisingly, however, overexpression of Hsp104 also cures [ PSI + ]. Several models have been proposed to explain this effect: inhibition of severing, asymmetric segregation of the seeds between mother and daughter cells, and dissolution of the prion seeds. First, we found that neither the kinetics of curing nor the heterogeneity in the distribution of the green fluorescent protein (GFP)-labeled Sup35 foci in partially cured yeast cells is compatible with Hsp104 overexpression curing [ PSI + ] by inhibiting severing. Second, we ruled out the asymmetric segregation model by showing that the extent of curing was essentially the same in mother and daughter cells and that the fluorescent foci did not distribute asymmetrically, but rather, there was marked loss of foci in both mother and daughter cells. These results suggest that Hsp104 overexpression cures [ PSI + ] by dissolution of the prion seeds in a two-step process. First, trimming of the prion seeds by Hsp104 reduces their size, and second, their amyloid core is eliminated, most likely by proteolysis.

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