Shorter Lag Phase Research Articles

EFFECT OF CRYOPRESERVATION ON GROWTH OF DIFFERENT ELITE EMBRYOGENIC CELL LINES OF NORWAY SPRUCE (PICEA ABIES (L.) KARST.)

Callus from seven different elite embryogenic cell lines of Norway spruce originated from the controlled crossing conducted by Skogforsk (Forest Research Institute of Sweden) have been cryopreserved in liquid nitrogen using the slow freezing method with sorbitol (0.2 and 0.4 M) and DMSO (5% v/v) as cryoprotectants. Cooloing of samples was performed in a programmable freezer Cryo Med 7452. The cryovials were inserted at +4°C and frozen at -0.3°C/min to -16°C. After 15 min at -16°C, to avoid rapid cooling and crystal formation in the cells, the cryovials were further cooled to -35°C at -0.3°C/min. At the end of the freezing program the cryovials were transferred and stored in a CBS V1500 liquid nitrogen storage unit at -196°C. After thawing and plating on proliferation medium, the recovered cells showed a short lag phase, after which they continued to grow. Lags in growth were also observed after the transfer to pre-maturation and maturation media. Somatic embryo maturation and plantlet regeneration occurred in all the tested embryogenic lines.

Rennet-Induced Aggregation of Heated pH-Adjusted Skim Milk

Heated (20-100 °C/0-30 min) skim milks (pH 6.5-7.1) were diluted in buffer (pH 7.0). Rennet was added, and the particle size with time was measured. For all samples, the size initially decreased (lag phase) and then increased (aggregation phase). Milks heated at ≤60 °C had short lag phases and rapid aggregation phases regardless of pH. Milks heated at >60 °C at pH 6.5 had long lag phases and slow aggregation phases. As the pH increased, the lag phase shortened and the aggregation phase accelerated. The aggregation time was correlated with the level of whey protein associated with the casein micelles and with the level of κ-casein dissociated from the micelles. Heated milks formed weak gels when renneted. It is proposed that the milks heated at low pH have whey proteins associated with the casein micelles and that these denatured whey proteins stabilize the micelles to aggregation by rennet and therefore inhibit gelation. In the milks heated at higher pH, the whey proteins associate with κ-casein in the serum and, on rennet treatment, the κ-casein-depleted micelles and the serum-phase whey protein/κ-casein complexes aggregate; however, the denatured whey proteins stabilize the aggregates so that gelation is still inhibited.

Nondestructive Evaluation of Quality Changes and the Optimum Time for Harvesting During Jujube (Zizyphus jujuba Mill. cv. Changhong) Fruits Development

Jujube fruit is an increasingly important fruit in the world for its high nutrients content and the harvest time is crucial for jujube fruit quality. The aim of the study was to evaluate the quality changes and the optimum time for harvesting of jujube fruit (Zizyphus jujuba Mill. cv. Changhong) by chlorophyll fluorescence. Our results showed that jujube fruit cv. Changhong displayed a double sigmoid growth curve with a very short lag phase. At the last three ripening stages (72, 80, and 88 days after petal fall), Fo, Fm, and Fv showed positive and significant correlation with DPPH radical scavenging activity, total phenolic, reducing sugar, ascorbic acid, and total flavonoid (0.729 ≤ r ≤ 0.920, P < 0.05). However, pH, total soluble sugar and carotenoids showed negative and significant correlation with Fo, Fm, and Fv (−0.885 ≤ r ≤ −0.826). The last three ripeness stages in our study were differentiated using the chlorophyll fluorescence parameters. Therefore, chlorophyll fluorescence is a helpful, nondestructive technique to evaluate the quality changes and the optimum time for harvesting of Changhong jujube.

Enhanced resistance of Saccharomyces cerevisiae to vanillin by expression of lacA from Trametes sp. AH28-2

Saccharomyces cerevisiae is affected by the presence of certain phenolic compounds such as vanillin during fermentation of pretreated lignocellulosic hydrolysates. Since vanillin can be polymerized in the presence of laccase into compounds with lower toxicity, the laccase gene, lacA, from Trametes sp. AH28-2 was fused to the α-factor signal sequence and transferred into S. cerevisiae CEN.PK strains for secretory expression. Furthermore, the chaperone gene, KAR2, was overexpressed to promote the translocation of laccase. In the presence of 8mmol/L vanillin, a shorter lag phase was observed in the lacA gene expressing strains. The vanillin-specific conversion rate of the lacA-expressing strain BSJX0A2 was 0.069gg−1biomassh−1, while it was 0.065gg−1biomassh−1 in the reference strain.

Pneumococcal replicative state in relation to its adherence capacity to A549-cell line: A preliminary in vitro analysis

This study was to compare the replication capacity of pneumococcal isolates (serotypes 1, 7F, 19F and 23F) with their adherence pattern to monolayer cells (A549). For standardization purposes, all isolates showed a normal growth curve in both bacteriological (THB + 0.5% yeast extract with and without 2% FBS) and cell culture media (RPMI + 2% FBS). In the former media, a shorter lag phase was observed for isolate serotypes 1 and 7F in presence of serum while in the later; growth yield was lower for all isolates with stationary phase approaching OD600 of 0.01 as compared to 1.0 in bacteriological media. In the replicative analysis at different growth phases of the isolates in cell culture media, growth capacity at 3 h post-incubation was frequently twice as that at 1 h, and that at early-log phase was frequently higher than that at mid-log phase at both post-incubation times. Adherence was frequently the least at early-log phase although the isolates were in the most active state of replication to increase the number of pneumococcal cells to adhere. At mid- and late-log phases, pneumococcal adherence was frequently higher although the replication was reduced. This study marks the potential correlation between pneumococcal growth fitness and adherence capacity whereby the later may not be superior during the early growth phase.

Effect of pig manure to grass silage ratio on methane production in batch anaerobic co-digestion of concentrated pig manure and grass silage

Anaerobic co-digestion of concentrated pig manure (PM) with grass silage (GS) at five different PM to GS volatile solid (VS) ratios of 1:0, 3:1, 1:1, 1:3 and 0:1 was evaluated by examining operation stability and methane (CH 4) production potentials. The highest specific CH 4 yields were 304.2 and 302.8 ml CH 4/g VS at PM to GS ratios of 3:1 and 1:1, respectively. The digestion systems failed at the ratio of 0:1. The lag phase lasted 29.5, 28.1, 24.6 and 21.3 days at the ratios of 1:0, 3:1, 1:1 and 1:3, respectively. The daily methane yield was linearly correlated with the acetic acid concentration, indicating methane production was probably associated with acetoclastic methanogenesis. The hydrolysis constant linearly decreased with increasing the fraction of GS in the feedstock. This study recommends applying the PM to GS ratio of 1:1 in practice due to a high specific methane yield and a short lag phase.

Virulence and pathogen multiplication: a serial passage experiment in the hypervirulent bacterial insect-pathogen Xenorhabdus nematophila.

The trade-off hypothesis proposes that the evolution of pathogens' virulence is shaped by a link between virulence and contagiousness. This link is often assumed to come from the fact that pathogens are contagious only if they can reach high parasitic load in the infected host. In this paper we present an experimental test of the hypothesis that selection on fast replication can affect virulence. In a serial passage experiment, we selected 80 lines of the bacterial insect-pathogen Xenorhabdus nematophila to multiply fast in an artificial culture medium. This selection resulted in shortened lag phase in our selected bacteria. We then injected these bacteria into insects and observed an increase in virulence. This could be taken as a sign that virulence in Xenorhabdus is linked to fast multiplication. But we found, among the selected lineages, either no link or a positive correlation between lag duration and virulence: the most virulent bacteria were the last to start multiplying. We then surveyed phenotypes that are under the control of the flhDC super regulon, which has been shown to be involved in Xenorhabdus virulence. We found that, in one treatment, the flhDC regulon has evolved rapidly, but that the changes we observed were not connected to virulence. All together, these results indicate that virulence is, in Xenorhabdus as in many other pathogens, a multifactorial trait. Being able to grow fast is one way to be virulent. But other ways exist which renders the evolution of virulence hard to predict.

Abnormal plasma fibrin clot characteristics are associated with worse clinical outcome in patients with peripheral arterial disease and thromboangiitis obliterans

BackgroundA role of blood coagulation in the pathogenesis of peripheral arterial disease (PAD) and Buerger's disease, or thromboangiitis obliterans (TAO), remains unclear. ObjectiveTo test the hypothesis that PAD and TAO are associated with prothrombotic phenotype of a fibrin clot. Patients and methodsEx vivo plasma fibrin clot permeability, turbidimetry and efficiency of fibrinolysis were investigated in 106 patients with PAD and 20 patients with TAO and compared with the respective control groups matched for age, sex, and cardiovascular risk factors. The progression of PAD and TAO were evaluated during follow-up of 3–7.5 years. PAD patients were characterized by lower clot permeability (−18.8%, p=0.005), shorter lag phase (−35.3%, p<0.001), higher maximum clot absorbancy (+22.4%, p<0.001), prolonged clot lysis time (+30.6%, p=0.003), and lower rate of D-dimer release from clots in the presence of recombinant tissue plasminogen activator (−16.5%, p=0.009), but twofold lower maximum D-dimer levels released from clots during lysis (p<0.001) than the controls. Similar, but more pronounced abnormalities were observed in TAO patients versus controls (all p<0.01). Seventeen PAD (16%) and 3 (15%) TAO patients were lost to follow-up. The progression observed in 47 (52.8%) PAD patients and 10 (59%) TAO patients was associated with lower clot permeability (−14.6%, p=0.009, and −17.5%, p=0.02) and prolonged clot lysis (+11.3%, p=0.004, and +12.4%, p=0.03, respectively). ConclusionsUnfavorably altered fibrin clot properties are observed in both PAD and TAO. Denser fibrin clots with reduced susceptibility to lysis might characterize the progression of both diseases during long-term follow-up.

Effect of caseinomacropeptide concentrate addition on the growth of bifidobacteria

We investigated the effect of caseinomacropeptide concentrate (CMP) on the growth and metabolic activity of Bifidobacterium bifidum CCDM 94 and Bifidobacterium lactis BB12 in skim milk with the addition of glucose and of various combinations of yeast extract, l-cysteine-HCl, ascorbic acid, and CMP. The microorganisms grew well on the medium containing a combination of all the nutrients mentioned, and on that where only l-cysteine-HCl&lt;br /&gt;was missing. In the media enhanced with CMP a shorter lag-phase occurred than in those without CMP. The shortest lag-phase and the longest log-phase with a high growth rate were observed in media No. 2 (complete medium with CMP) and No. 8 (complete medium without l-cysteine-HCl).

Sucrose Transfructosylation to Fructooligosaccharides by the Application of Commercial Enzymes

and water. Time course of CO2 production in each trial was fitted employing theDMFit software, toestimate lagphaseduration (min) and CO2 production rate ( max, min-1); R2 obtained were in the range 0.998311 – 0.999478 and s.e. fit 1.750 – 2.891. Results highlighted that in dough systems Z. mobilis and S. cerevisiaewere found toproduce similar amount of CO2, 187 and212ml CO2/g dcw respectively, after 120min incubation. Time course of CO2 production in S. cerevisiae was characterized by a short lag phase duration (9.7min) and 2.2min-1 max. CO2 production by Z. mobiliswas instead characterized by a longer lag phase (30.3min), that increased up to 56min after 12 d cell storage, but a higher max (2.7min-1). From an overall look at the obtained results it can be concluded that Z. mobilis can be proposed as valid alternative to S. cerevisiae to be employed as leavening agent in baked-products.

Phenol biodegradation by the thermoacidophilic archaeon Sulfolobus solfataricus 98/2 in a fed-batch bioreactor

Toxic at low concentrations, phenol is one of the most common organic pollutants in air and water. In this work, phenol biodegradation was studied in extreme conditions (80°C, pH = 3.2) in a 2.7 l bioreactor with the thermoacidophilic archaeon Sulfolobus solfataricus 98/2. The strain was first acclimatized to phenol on a mixture of glucose (2000 mg l(-1)) and phenol (94 mg l(-1)) at a constant dissolved oxygen concentration of 1.5 mg l(-1). After a short lag-phase, only glucose was consumed. Phenol degradation then began while glucose was still present in the reactor. When glucose was exhausted, phenol was used for respiration and then for biomass build-up. After several batch runs (phenol < 365 mg l(-1)), specific growth rate (μ(X)) was 0.034 ± 0.001 h(-1), specific phenol degradation rate (q(P)) was 57.5 ± 2 mg g(-1) h(-1), biomass yield (Y(X/P)) was 52.2 ± 1.1 g mol(-1), and oxygen yield factor (Y(X/O2)) was 9.2 ± 0.2 g mol(-1). A carbon recovery close to 100% suggested that phenol was exclusively transformed into biomass (35%) and CO(2) (65%). Molar phenol oxidation constant (Y(O2/P)) was calculated from stoichiometry of phenol oxidation and introducing experimental biomass and CO(2) conversion yields on phenol, leading to values varying between 4.78 and 5.22 mol mol(-1). Respiratory quotient was about 0.84 mol mol(-1), very close to theoretical value (0.87 mol mol(-1)). Carbon dioxide production, oxygen demand and redox potential, monitored on-line, were good indicators of growth, substrate consumption and exhaustion, and can therefore be usefully employed for industrial phenol bioremediation in extreme environments.

Deacclimation kinetics and carbohydrate changes in stem tissues of Hydrangea in response to an experimental warm spell

Temperate winters are becoming progressively milder due to climate warming, and temperature patterns are becoming increasingly irregular with risk of unseasonable warm spells. Warm spells may cause premature loss of plant cold hardiness and increase the risk of subsequent freezing injury. This study investigated the timing and rate of deacclimation and associated changes in soluble carbohydrates and water status in stems of Hydrangea macrophylla ssp. macrophylla (Thunb.) Ser. ‘Alma’ and Hydrangea paniculata Sieb. ‘Vanille Fraise’ in response to a simulated warm spell (22°C/17°C day/night). In H. macrophylla, deacclimation kinetics showed a sigmoid course with a short lag-phase followed by a fast deacclimation rate. In H. paniculata, the deacclimation pattern could not be determined precisely, but H. paniculata, the hardier genotype based on mid-winter freezing tolerance, deacclimated to a greater extent than H. macrophylla. These results imply that dehardening resistance is not related to mid-winter hardiness. In both species deacclimation was associated with rehydration and decreasing sugar levels, but species-specific quantitative and qualitative differences in the accumulation patterns of specific sugars were observed. In H. paniculata cold hardiness may be associated with 1-kestose, an oligofructan frequently associated with overwintering in herbaceous plants, but not previously related to freezing tolerance in woody perennials.

Tobacco biomass hydrolysate enhances coenzyme Q10 production using photosynthetic Rhodospirillum rubrum

Coenzyme Q10 (CoQ10), a potent antioxidative dietary supplement, was produced using a photosynthetic bacteria Rhodospirillum rubrum ATCC 25852 by submerged fermentation supplemented with tobacco biomass hydrolysate (TBH) in comparison with media supplemented with hydrolysates prepared with alfalfa (ABH) or spinach (SBH). Growth medium supplemented with 20% (v/v) TBH was found favorable with regard to cell density and CoQ10 concentration. The stimulation effects on cell growth (shortened lag phase, accelerated exponential growth, and elevated final cell concentration) and CoQ10 production (enhanced specific CoQ10 content per unit cell weight) could be attributed to the presence of solanesol, the precursor of CoQ10, in the tobacco biomass. The final yield of CoQ10 reached 20.16 mg/l in the fermentation medium supplemented with 20% TBH.

Exploring the lag phase and growth initiation of a yeast culture by means of an individual-based model

The performance of fermentation processes is greatly influenced by the size and quality of inocula. The characterization of the replicative age is decided by the number of birth scars each yeast exhibits on its cellular membrane. Yeast ageing and inoculum size are factors that affect industrial fermentation, particularly those processes in which the yeast cells are reused such as the production of beer. This process reuses yeast cropped at the end of one fermentation in the following one, in a process called “serial repitching”. The aim of this study was to explore the effects of inoculum size and ageing on the first stages of the dynamics of yeast population growth. However, only Individual-based Models (IbMs) allow the study of small, well-characterized, microbial inocula. We used INDISIM-YEAST, based on the generic IbM simulator INDISIM, to carry out these studies. Several simulations were performed to analyze the effect of the inoculum size and genealogical age of the cells that made it up on the lag phase, first division time and specific growth rate. The shortest lag phase and time to the first division were obtained with largest inocula and with the youngest inoculated parent cells.

A water–sediment screening tool for measuring biodegradation of organic chemicals

A water–sediment screening tool (WSST) was developed based on OECD guideline 301 C (MITI I; Ministry of International Trade and Industry, Japan) to generate biodegradation data. The WSST and experimental procedures were tested and validated using aniline (CAS No. 62-53-3) and benzoic acid (CAS No. 65-85-0) as reference substances. In the presence of sediment components a higher endogenous respiration rate in the control vessels without test substance was measured compared to the water-only MITI test system, particularly due to organic constituents. However, it could be demonstrated that a distinct biodegradation in the presence of sediment can be determined and that there is no influence of the sediment pre-treatment on the biological oxygen demand in the WSST. Experiments resulted in biodegradation rates > 60% after approximately six days for both compounds. However, degradation of benzoic acid resulted in a shorter lag-phase and a higher degree of degradation compared to aniline. Differences in results between the MITI test system and the WSST observed for aniline can be explained by adsorption to constituents of the sediment and assimilation by activated sludge. In comparison with literature data the results obtained for aniline in the MITI test system and the WSST showed reproducibility and were within the expected range. In conclusion, the WSST is a suitable screening tool to determine kinetic biodegradation data required to predict the biodegradation behaviour of organic chemicals in water–sediment systems and the data might be used to improve quantitative structure–property relationships (QSPRs).

Transport and degradation of metalaxyl and isoproturon in biopurification columns inoculated with pesticide-primed material

Laboratory column displacement experiments were performed to examine whether addition of pesticide-primed material to the matrix of an on-farm biopurification system (BPS), intended to remove pesticides from agricultural waste water, positively affects the degradation of mobile pesticides in the system. Percolated column microcosms with varying types and amounts of metalaxyl and/or isoproturon-primed material or non-primed material were irrigated with water artificially contaminated with isoproturon and/or metalaxyl. Transport of isoproturon was well described using the convection dispersion equation and no dissipation was observed, even in columns inoculated with isoproturon-primed material. On the other hand, delayed dissipation of metalaxyl, i.e., after an initial lag phase, was encountered in all columns receiving metalaxyl. In all systems, dissipation could be described using the Monod model indicating that a metalaxyl degrading population grew in the systems. There was a clear correlation between the lag phase and the amount of metalaxyl-primed material added to the system, i.e., increasing amounts of added material resulted into shorter lag phases and hence more rapid initiation of growth-associated metalaxyl degradation in the system. Our observations suggest that indeed pesticide-primed material can reduce the start-up phase of degradation of mobile pesticides in a BPS and as such can increase its efficiency. However, the primed material should be chosen carefully and preferentially beforehand tested for its capacity to degrade the pesticide.

Lignin and starch as potential inductors for biodegradation of films based on poly(vinyl alcohol) and protein hydrolysate

The objective of this work was to study the biodegradation of blow-moulded films of poly(vinyl alcohol) (PVA)/protein hydrolysate (PH) which contain biodegradation inductors of the starch (S) and lignin (LI) types. These increase the biodegradation rate of PVA while preserving or improving the technical and usage properties of blends. The aim of the work was to reach the maximum breakdown rate so that rapid disintegration of PVA could take place at a wastewater treatment plant. The biological material chosen was activated sludge from a municipal wastewater treatment plant. Preparation of the blends required that a plasticiser be used, in this case glycerol (G). This allowed for successful processing but prolonged the lag phase of PVA breakdown as well as reducing its final biodegradation percentage. The influence of G, in this respect, was not affected by incorporating PH. S and LI reduced the influence of the plasticiser but caused a breakdown rate comparable to PVA itself. Contrarily, after adapting biomass to PVA, applying G produced a PVA breakdown rate three times greater, albeit with a lag phase prolonged fivefold. However, due to the duration of breakdown (the period above the retention time of wastewater during activation), this effect was not positive. The addition of PH to the blends mentioned did not exhibit any clear favourable influence. Adding S resulted in a shorter lag phase, in addition to which the degradation rate increased by approximately 1.5 times. Combining LI and S distinctly accelerated the degradation of a blend, although a disadvantage of doing so is an incomplete breakdown of the substrate, which lowers the final biodegradation percentage. Therefore, an eventual compromise was arrived at, this being a blend of PVA/G PH S. Its breakdown time is half that of pure PVA, and the films produced, from a mechanical standpoint, are more convenient.

Listeria monocytogenes Attachment to and Detachment from Stainless Steel Surfaces in a Simulated Dairy Processing Environment

The presence of pathogens in dairy products is often associated with contamination via bacteria attached to food-processing equipment, especially from areas where cleaning/sanitation is difficult. In this study, the attachment of Listeria monocytogenes on stainless steel (SS), followed by detachment and growth in foods, was evaluated under conditions simulating a dairy processing environment. Initially, SS coupons were immersed in milk, vanilla custard, and yogurt inoculated with the pathogen (10(7) CFU/ml or CFU/g) and incubated at two temperatures (5 and 20 degrees C) for 7 days. By the end of incubation, cells were mechanically detached from coupons and used to inoculate freshly pasteurized milk which was subsequently stored at 5 degrees C for 20 days. The suspended cells in all three products in which SS coupons were immersed were also used to inoculate freshly pasteurized milk (5 degrees C for 20 days). When SS coupons were immersed in milk, shorter lag phases were obtained for detached than for planktonically grown cells, regardless of the preincubation temperature (5 or 20 degrees C). The opposite was observed when custard incubated at 20 degrees C was used to prepare the two types of inocula. However, in this case, a significant increase in growth rate was also evident when the inoculum was derived from detached cells. In another parallel study, while L. monocytogenes was not detectable on SS coupons after 7 days of incubation (at 5 degrees C) in inoculated yogurt, marked detachment and growth were observed when these coupons were subsequently transferred and incubated at 5 degrees C in fresh milk or/and custard. Overall, the results obtained extend our knowledge on the risk related to contamination of dairy products with detached L. monocytogenes cells.

Efficient phenol degradation by a newly characterized Pseudomonas sp. SA01 isolated from pharmaceutical wastewaters

Using the phenol enrichment cultivation method, we succeeded in screening a few bacterial strains with a varied ability for utilizing phenol as the sole carbon source from pharmaceutical disposal wastewaters plant. Among these strains, a potent phenol-degrading bacterium, assigned Pseudomonas sp. SA01, was identified and examined as having the greatest potential for degrading phenol. According to biochemical characteristics and 16S rRNA sequence analysis, the isolate was identified as Pseudomonas sp. The isolated strain started to degrade 0.7 g/l of phenol after an initial very short lag phase, and phenol decomposition was then rapidly completed within 30 h. Pseudomonas sp. SA01 was able to degrade phenol in concentrations up to 1 g/l. Higher phenol concentrations (>1 g/l) had a significant inhibitory effect on bacterial growth. The optimum degradation pH value was found to be 6.5. Addition of mannitol and casein as auxiliary carbon and nitrogen sources enhanced the rate of phenol removal to as low as 20 h. Based on the absorption spectra of catechol bioconversion of phenol-grown cells, it was concluded that the SA01 strain metabolizes the phenol via a meta-cleavage pathway.

Competitive biodegradation of dichlobenil and atrazine coexisting in soil amended with a char and citrate

The role of char nutrients in the biodegradation of coexisting dichlobenil and atrazine in a soil by their respective bacterial degraders, DDN and ADP, was evaluated. Under growing conditions, their degradation in soil extract was slow with <40% and <20% degraded within 64 h, respectively. The degradation in extracts and slurries of char-amended solids increased with increasing char content, due to nutritional stimulation on microbial activities. By supplementing soil extract with various major nutrients, the measured degradation demonstrated that P was the exclusive limiting nutrient. The reduction in the degradation of coexisting dichlobenil and atrazine resulted apparently from the competitive utilization of P by DDN and ADP. With a shorter lag phase, ADP commenced growing earlier than DDN with the advantage of utilizing P first in insufficient supply. This resulted in an inhibition on the growth of DDN and thus suppression on dichlobenil degradation.