Transcriptome analysis and physiology of Bifidobacterium longum NCC2705 cells under continuous culture conditions

2012 ◽  
Vol 3 (4) ◽  
pp. 261-272 ◽  
Author(s):  
V. Mozzetti ◽  
F. Grattepanche ◽  
D. Moine ◽  
B. Berger ◽  
E. Rezzonico ◽  
...  
Keyword(s):  
Continuous Culture ◽  
Bifidobacterium Longum ◽  
Viable Cell ◽  
Culture Conditions ◽  
Stress Conditions ◽  
Stress Factors ◽  
Bacterial Cells ◽  
Continuous Cultures ◽  
Batch Cultures ◽  
Cell Counts

A central issue in the use of probiotics in food and food supplements is their sensitivity to many environmental stress factors. The resistance of probiotic cells to lethal stress can be improved by application of homologous or heterologous sub-lethal stress during culture. This screening procedure is generally performed using batch cultures. Continuous cultures could be a suitable and more efficient method to test different stress factors on one culture instead of repeating several batch cultures. However, before testing stresses using continuous cultures, the physiological stability of continuously produced cells over a considered time period must be first evaluated. A continuous culture of Bifidobacterium longum NCC2705 was maintained for 211 h at a dilution rate of 0.1 per h, mimicking a deceleration growth phase culture. Stable viable cell counts were measured over the culture period, decreasing only moderately from 8.8 to 8.6 log10 cfu/ml. A slight shift in metabolite production, characterized by increased lactate and decreased acetate, formate and ethanol concentrations was observed. Susceptibilities to antibiotics and stress conditions were stable (cefotaxim, ampicillin, ceftazidime) or moderately affected (simulated gastric juices, heat, bile salts, tetracycline, chloramphenicol, penicillin, vancomycin and neomycin) over culturing time. Comparison of gene transcription profiles between samples collected after 31 h of continuous culture and samples collected after 134 and 211 h revealed only limited changes in expression of 1.0 and 3.8% of total genes, respectively. Based on these results, we propose that continuous culture can be used to produce bacterial cells with stable physiological properties suitable for fast and efficient screening of sub-lethal stress conditions.

Fast screening of Bifidobacterium longum sublethal stress conditions in a novel two-stage continuous culture strategy

2013 ◽  
Vol 4 (2) ◽  
pp. 167-178 ◽  
Author(s):  
V. Mozzetti ◽  
F. Grattepanche ◽  
B. Berger ◽  
E. Rezzonico ◽  
F. Arigoni ◽  
...  
Keyword(s):  
Cell Viability ◽  
Continuous Culture ◽  
Bifidobacterium Longum ◽  
Food Additives ◽  
Stress Conditions ◽  
Stress Factors ◽  
Exponential Growth Phase ◽  
Two Stage ◽  
Batch Cultures ◽  
Sublethal Stress

A central issue in the application of probiotics as food additives is their fastidious production and their sensitivity to many environmental stresses. The importance of inducible cell-protective mechanisms triggered by application of sublethal stresses for survival under stress conditions has been demonstrated. Continuous cultures could be a suitable and more efficient method to test stress factors on one culture instead of several repeated batch cultures. In this study, the application of a two-stage continuous culture of Bifidobacterium longum NCC2705 was investigated. The first reactor was operated under fixed conditions at 37 °C and pH 6.0 and used to produce cells with controlled physiology, mimicking cells in the late exponential growth phase. Stress pretreatment combinations of pH (6.0, 5.0 and 4.0), temperature (37, 45 and 47 °C) and NaCl (0, 5 and 10%) were tested in the second reactor. Of all tested combinations, only those of pH 4.0 significantly decreased cell viability in the second reactor compared to control conditions (37 °C, pH 6.0, 0% NaCl) and, therefore, could not be considered as sublethal stresses. Pretreatments with 5 or 10% NaCl had a negative effect on cell viability after gastric lethal stress. A significant improvement in cell resistance to heat lethal stress (56 °C, 5 min) was observed for cells pretreated at 47 °C. In contrast, heat pretreatment negatively affected cell viability after freeze drying and osmotic lethal stresses. The two-stage continuous culture allowed for efficient screening of several stress pretreatments during the same experiment with up to four different conditions tested per day. Optimal sublethal stress conditions can also be applied for producing cells with traditional batch cultures.

Effect of carbon dioxide on growth of Pseudomonas putida ATCC 11172 on asparagine, citrate, glucose, and lactate in batch and continuous culture

1985 ◽  
Vol 31 (9) ◽  
pp. 763-766 ◽  
Author(s):  
Göran Molin
Keyword(s):  
Carbon Dioxide ◽  
Growth Rate ◽  
Pseudomonas Putida ◽  
Continuous Culture ◽  
Specific Growth ◽  
Carbon Sources ◽  
Maximum Specific Growth Rate ◽  
The Other ◽  
Continuous Cultures ◽  
Batch Cultures

The growth of Pseudomonas putida ATCC 11172 on L-asparagine, citrate, D-glucose, and L-lactate was followed in air and in 40% CO2 + air, using batch and carbon-limited continuous cultures. Batch cultures in air utilized a mixture of the carbon sources simultaneously. However, a change to 40% CO2 favoured the utilization of glucose. The maximum specific growth rate (μmax) in air was about 0.3 h−1 on glucose and 0.6 h−1 on the other carbon sources. In CO2, the μmax for glucose was reduced by 16% compared with almost 60–70% for the others. An order of preference for the different carbon sources in continuous cultures was determined by comparing the dilution rates at which the different carbon sources started to appear in the effluent. Glucose was the first compound to appear as the dilution rate increased (lowest preference when grown in air). In 40% CO2, the μmax for glucose was slightly higher than the others and the recorded preference for glucose in continuous culture was equal to that for citrate but was somewhat lower than that of lactate and asparagine. D-Gluconate and glucono-δ-lactone were produced as a step in the utilization of glucose. The D-gluconate production was enhanced by CO2.

scholarly journals Bactericidal Activity of ACH-702 against Nondividing and Biofilm Staphylococci

2012 ◽  
Vol 56 (7) ◽  
pp. 3812-3818 ◽  
Author(s):  
Steven D. Podos ◽  
Jane A. Thanassi ◽  
Melissa Leggio ◽  
Michael J. Pucci
Keyword(s):  
Protein Synthesis ◽  
Stationary Phase ◽  
Bactericidal Activity ◽  
Bacterial Infections ◽  
De Novo ◽  
Viable Cell ◽  
Bacterial Cells ◽  
Protein Synthesis Inhibitors ◽  
Content Type ◽  
Cell Counts

ABSTRACTMany bacterial infections involve slow or nondividing bacterial growth states and localized high cell densities. Antibiotics with demonstrated bactericidal activity rarely remain bactericidal at therapeutic concentrations under these conditions. The isothiazoloquinolone (ITQ) ACH-702 is a potent, bactericidal compound with activity against many antibiotic-resistant pathogens, including methicillin-resistantStaphylococcus aureus(MRSA). We evaluated its bactericidal activity under conditions where bacterial cells were not dividing and/or had slowed their growth. AgainstS. aureuscultures in stationary phase, ACH-702 showed concentration-dependent bactericidal activity and achieved a 3-log-unit reduction in viable cell counts within 6 h of treatment at ≥16× MIC values; in comparison, the bactericidal quinolone moxifloxacin and the additional comparator compounds vancomycin, linezolid, and rifampin at 16× to 32× MICs showed little or no bactericidal activity against stationary-phase cells. ACH-702 at 32× MIC retained bactericidal activity against stationary-phaseS. aureusacross a range of inoculum densities. ACH-702 did not kill cold-arrested cells yet remained bactericidal against cells arrested by protein synthesis inhibitors, suggesting that its bactericidal activity against nondividing cells requires active metabolism but notde novoprotein synthesis. ACH-702 also showed a degree of bactericidal activity at 16× MIC againstS. epidermidisbiofilm cells that was superior to that of moxifloxacin, rifampin, and vancomycin. The bactericidal activity of ACH-702 against stationary-phase staphylococci and biofilms suggests potential clinical utility in infections containing cells in these physiological states.

The species-specific effects of sublethal concentrations of cadmium on freshwater phytoplankton communities in a Canadian Shield lake

1985 ◽  
Vol 63 (11) ◽  
pp. 1997-2003 ◽  
Author(s):  
Danny C. Reinke ◽  
Frank DeNoyelles Jr.
Keyword(s):  
Cadmium Concentration ◽  
Continuous Cultures ◽  
Experimental Lakes Area ◽  
Batch Cultures ◽  
Freshwater Phytoplankton ◽  
Specific Effects ◽  
Cell Counts ◽  
Phytoplankton Communities ◽  
Species Specific

The species-specific responses of natural phytoplankton communities to low cadmium concentrations were measured in Lake 239 (Experimental Lakes Area, northwestern Ontario). Both in situ and laboratory 5-L continuous-flow cultures, and 5-L and 100-mL cultures were used. Asterionella formosa, Dinobryon sertularia, and Dinobryon bavaricum showed dramatic negative sensitivity to low cadmium concentrations (5–100 μg/L), while Rhabdoderma gorskii and Elakatothrix sp. consistently increased in numbers at the same cadmium concentrations. In all experiments, some species exhibited no apparent effect to cadmium addition as measured by cell counts. The "bottle effect" of each technique was evaluated by comparing the community similarity valves of the control cultures to the lake samples and showed the in situ continuous cultures to be most similar to the lake followed by the laboratory continuous cultures, the in situ 5-L batch cultures, the 5-L laboratory cultures, and the 100-mL batch cultures. Replicate cadmium cultures, all techniques, were more similar to each other than the lake samples. The similarity of the cadmium cultures to the lake sample or control cultures decreased with increased cadmium concentration and incubation time.

scholarly journals Cellular Responses to Postsegregational Killing by Restriction-Modification Genes

2000 ◽  
Vol 182 (8) ◽  
pp. 2218-2229 ◽  
Author(s):  
Naofumi Handa ◽  
Asao Ichige ◽  
Kohji Kusano ◽  
Ichizo Kobayashi
Keyword(s):  
Viable Cell ◽  
Underlying Mechanism ◽  
Temperature Sensitive ◽  
Dependent Manner ◽  
Bacterial Cells ◽  
Gene Complex ◽  
Cellular Processes ◽  
Cell Counts ◽  
Gene Complexes ◽  
Restriction Modification

ABSTRACT Plasmids that carry one of several type II restriction modification gene complexes are known to show increased stability. The underlying mechanism was proposed to be the lethal attack by restriction enzyme at chromosomal recognition sites in cells that had lost the restriction modification gene complex. In order to examine bacterial responses to this postsegregational cell killing, we analyzed the cellular processes following loss of the EcoRI restriction modification gene complex carried by a temperature-sensitive plasmid in anEscherichia coli strain that is wild type with respect to DNA repair. A shift to the nonpermissive temperature blocked plasmid replication, reduced the increase in viable cell counts and resulted in loss of cell viability. Many cells formed long filaments, some of which were multinucleated and others anucleated. In a mutant defective in RecBCD exonuclease/recombinase, these cell death symptoms were more severe and cleaved chromosomes accumulated. Growth inhibition was also more severe in recA, ruvAB, ruvC,recG, and recN mutants. The cells induced the SOS response in a RecBC-dependent manner. These observations strongly suggest that bacterial cells die as a result of chromosome cleavage after loss of a restriction modification gene complex and that the bacterial RecBCD/RecA machinery helps the cells to survive, at least to some extent, by repairing the cleaved chromosomes. These and previous results have led us to hypothesize that the RecBCD/Chi/RecA system serves to destroy restricted “nonself” DNA and repair restricted “self” DNA.

scholarly journals The Joint Effect of pH Gradient and Glucose Feeding on the Growth Kinetics of Lactococcus lactis CECT 539 in Glucose-Limited Fed-Batch Cultures

2019 ◽  
Vol 68 (2) ◽  
pp. 269-280
Author(s):  
MÓNICA COSTAS MALVIDO ◽  
ELISA ALONSO GONZÁLEZ ◽  
RICARDO J. BENDAÑA JÁCOME ◽  
NELSON PÉREZ GUERRA
Keyword(s):  
Lactococcus Lactis ◽  
Viable Cell ◽  
Product Formation ◽  
Ph Gradient ◽  
Feeding Strategies ◽  
Fed Batch ◽  
Batch Cultures ◽  
Cell Counts ◽  
First Time ◽  
Kinetics Of

Two glucose-limited realkalized fed-batch cultures of Lactococcus lactis CECT 539 were carried out in a diluted whey medium (DW) using two different feeding media. The cultures were fed a mixture of a 400 g/l concentrated lactose and a concentrated mussel processing waste (CMPW, 101.72 g glucose/l) medium (fermentation I) or a CMPW medium supplemented with glucose and KH2PO4 up to concentrations of 400 g glucose/l and 3.21 g total phosphorus/l, respectively (fermentation II). For an accurate description and a better understanding of the kinetics of both cultures, the growth and product formation by L. lactis CECT 539 were both modelled, for the first time, as a function of the amounts of glucose (G) added and the pH gradient (VpH) generated in every realkalization and feeding cycle, by using an empirical polynomial model. With this modeling procedure, the kinetics of biomass, viable cell counts, nisin, lactic acid, acetic acid and butane-2,3-diol production in both cultures were successfully described (R2 values > 0.970) and interpreted for the first time. In addition, the optimum VpH and G values for each product were accurately calculated in the two realkalized fed-batch cultures. This approach appears to be useful for designing feeding strategies to enhance the productions of biomass, bacteriocin, and metabolites by the nisin-producing strain in wastes from the food industry.

scholarly journals Microencapsulation of Lactococcus lactis Gh1 with Gum Arabic and Synsepalum dulcificum via Spray Drying for Potential Inclusion in Functional Yogurt

Molecules ◽  
2019 ◽  
Vol 24 (7) ◽  
pp. 1422 ◽  
Author(s):  
Nurul Farhana Fazilah ◽  
Nurmelissa Hanani Hamidon ◽  
Arbakariya B. Ariff ◽  
Mohd Ezuan Khayat ◽  
Helmi Wasoh ◽  
...  
Keyword(s):  
Spray Drying ◽  
Lactococcus Lactis ◽  
Gum Arabic ◽  
Streptococcus Thermophilus ◽  
Viable Cell ◽  
Starter Cultures ◽  
Lactobacillus Delbrueckii ◽  
Bacterial Cells ◽  
Cell Counts

There has been an explosion of probiotic incorporated based product. However, many reports indicated that most of the probiotics have failed to survive in high quantity, which has limited their effectiveness in most functional foods. Thus, to overcome this problem, microencapsulation is considered to be a promising process. In this study, Lactococcus lactis Gh1 was encapsulated via spray-drying with gum Arabic together with Synsepalum dulcificum or commonly known as miracle fruit. It was observed that after spray-drying, high viability (~109 CFU/mL) powders containing L. lactis in combination with S. dulcificum were developed, which was then formulated into yogurt. The tolerance of encapsulated bacterial cells in simulated gastric juice at pH 1.5 was tested in an in-vitro model and the result showed that after 2 h, cell viability remained high at 1.11 × 106 CFU/mL. Incubation of encapsulated cells in the presence of 0.6% (w/v) bile salts showed it was able to survive (~104 CFU/mL) after 2 h. Microencapsulated L. lactis retained a higher viability, at ~107 CFU/mL, when incorporated into yogurt compared to non-microencapsulated cells ~105 CFU/mL. The fortification of microencapsulated and non-microencapsulated L. lactis in yogurts influenced the viable cell counts of yogurt starter cultures, Lactobacillus delbrueckii subs. bulgaricus and Streptococcus thermophilus.

scholarly journals Persister cells mediate tolerance to metal oxyanions in Escherichia coli

Microbiology ◽  
2005 ◽  
Vol 151 (10) ◽  
pp. 3181-3195 ◽  
Author(s):  
Joe J. Harrison ◽  
Howard Ceri ◽  
Nicole J. Roper ◽  
Erin A. Badry ◽  
Kimberley M. Sproule ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Viable Cell ◽  
Small Population ◽  
Water Soluble ◽  
Bacterial Cells ◽  
Persister Cells ◽  
Term Survival ◽  
E Coli ◽  
Cell Counts ◽  
High Concentrations

Bacterial cultures produce subpopulations of cells termed ‘persisters’, reputedly known for high tolerance to killing by antibiotics. Ecologically, antibiotics produced by competing microflora are only one potential stress encountered by bacteria. Another pressure in the environment is toxic metals that are distributed ubiquitously by human pollution, volcanic activity and the weathering of minerals. This study evaluated the time- and concentration-dependent killing of Escherichia coli planktonic and biofilm cultures by the water-soluble metal(loid) oxyanions chromate (), arsenate (), arsenite (), selenite (), tellurate () and tellurite (). Correlative to previous reports in the literature, control antibiotic assays indicated that a small proportion of E. coli biofilm populations remained recalcitrant to killing by antibiotics (even with 24 h exposure). In contrast, metal oxyanions presented a slow, bactericidal action that eradicated biofilms. When exposed for 2 h, biofilms were up to 310 times more tolerant to killing by metal oxyanions than corresponding planktonic cultures. However, by 24 h, planktonic cells and biofilms were eradicated at approximately the same concentration in all instances. Coloured complexes of metals and chelators could not be generated in biofilms exposed to or , suggesting that the extracellular polymeric matrix of E. coli may have a low binding affinity for metal oxyanions. Viable cell counts at 2 and 24 h exposure revealed that, at high concentrations, all of the metal oxyanions had killed 99 % (or a greater proportion) of the bacterial cells in biofilm populations. It is suggested here that the short-term survival of <1 % of the bacterial population corresponds well with the hypothesis that a small population of persister cells may be responsible for the time-dependent tolerance of E. coli biofilms to high concentrations of metal oxyanions.

Combined Effect of Water Activity and pH on the Inhibition of Escherichia coli by Nisin

2001 ◽  
Vol 64 (10) ◽  
pp. 1510-1514 ◽  
Author(s):  
PATRICIA CERRUTTI ◽  
MAURICIO R. TEREBIZNIK ◽  
MARTA SEGOVIA de HUERGO ◽  
ROSA JAGUS ◽  
ANA M. R. PILOSOF
Keyword(s):  
Escherichia Coli ◽  
Water Activity ◽  
Viable Cell ◽  
Stress Factors ◽  
Doehlert Design ◽  
Survival Fraction ◽  
E Coli ◽  
Cell Counts ◽  
Maximum Inhibition ◽  
Influence Of Ph

The Doehlert design and surface response methodology were used to study the influence of pH and water activity (aw) on Escherichia coli inhibition by nisin. Combining stress factors at levels where they are not inhibitory by themselves, a reduction of E. coli survival fraction can be achieved with lower nisin doses than in a single nisin treatment. For all the pH values assayed, a synergistic effect of aw and nisin concentration was detected, and the isoresponse lines showed the existence of an area of maximum inhibition. Factors that reduced viable cell counts by 4 to 5 log cycles were 1,000 to 1,400 IU of nisin per ml at pH 5.5 to 6.5 and a water activity of 0.97 and 0.98. The addition of different ionic and nonionic solutes to control aw suggested that the effect of aw in the inhibitory action of nisin on E. coli cells was not solute-specific. The use of the Doehlert experimental design was effective to determine the optimal combination of stress factors, as well as to point out the most important variables that affected E. coli inhibition.

scholarly journals Tolerance of Lactobacillus sakei to Osmotic Stress

2021 ◽  
pp. 574-583
Author(s):  
Anna Nikiforova ◽  
Sofia Khazagaeva ◽  
Irina Khamagaeva
Keyword(s):  
Lactic Acid ◽  
Sodium Chloride ◽  
Lactic Acid Bacteria ◽  
Osmotic Stress ◽  
New Technologies ◽  
Viable Cell ◽  
Inhibitory Effect ◽  
Bacterial Cells ◽  
Cell Counts ◽  
Nacl Concentration

Introduction. The development of new technologies of bacterial cultures for fish products is an important area of re-search. Sodium chloride (NaCl) has an inhibitory effect on most microorganisms and is often used in fish industry. The present research objective was to study the effect of NaCl on the growth of lactic acid bacteria of the species Lactoba-cillus sakei. Study objects and methods. The research featured strains of lactic acid bacteria L. sakei (L. sakei LSK-45 and L. sakei DSM 20017). The bacteria were cultivated in a semi-liquid MRS medium supplemented with sodium chloride at vari-ous concentrations. The optical density (OD) of the medium and the viable cell counts served as indicators of bacterial growth. The morphology of bacterial cells was studied by microscopic examination of Gram-stained bacteria. Results and discussion. Both L. sakei strains proved tolerant to NaCl concentrations up to 6% as proved by the high density of the cell population – 109 CFU/cm3. When NaCl concentration reached 10%, L. sakei LSK-45 demonstrated the highest resistance to osmotic stress. At the end of cultivation, the population density dropped to 108 CFU/cm3, and the survival rate was 92%. The number of viable cells of L. sakei DSM 20017 decreased to 106 CFU/cm3, and the sur-vival rate reached 62%. The high resistance of L. sakei LSK-45 to osmotic stress was caused by cell cohesion, which increased with the growing NaCl concentration and was regulated by the quorum sensing system. Conclusion. L. sakei showed a flexible response to the changes in NaCl concentration. L. sakei LSK-45 strain had the highest resistance to osmotic stress.

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