Glutamate dehydrogenase activity in lactobacilli and the use of glutamate dehydrogenase-producing adjunct Lactobacillus spp. cultures in the manufacture of cheddar cheese

AIMS: The study was undertaken to investigate the occurrence of glutamate dehydrogenase activity in different species of lactobacilli, and to determine, in a series of cheese-making trials, the effects of glutamate dehydrogenase-producing adjunct cultures on sensory attribute development during the maturation of cheddar cheese. METHODS AND RESULTS: The presence of dehydrogenase activity with glutamate as substrate was monitored in cell lysates of >100 strains from 30 different species of lactobacilli using a qualitative colorimetric plate screening assay. Activity was detectable in 25 of the 29 representative species obtained from culture collections and in 12 of the 13 non-starter species isolated from cheese. There were pronounced interspecies and strain differences in the occurrence, level and pyridine nucleotide specificity of the glutamate dehydrogenase activity detected. Among the non-starter lactobacilli the highest frequency of enzyme occurrence and activity was detected in the Lactobacillus plantarum isolates. The establishment of glutamate dehydrogenase-producing adjunct strains in the predominant population of lactobacilli in the cheese curd affected the formation of a number of volatile compounds in ripening cheddar cheese, while the presence of Lact. plantarum strains, in particular, was associated with an intensification and acceleration of aroma and flavour development during the maturation period. CONCLUSIONS: Glutamate dehydrogenase formation by lactobacilli is a strain-dependent metabolic attribute, and adjunct cultures expressing the activity that are able to proliferate during cheese ripening have a positive impact on the rate of development and the intensity of cheddar cheese aroma and flavour development. SIGNIFICANCE AND IMPACT OF THE STUDY: It has been demonstrated that some strains of glutamate dehydrogenase-producing lactobacilli have potential use as adjunct cultures to accelerate and intensify aroma and flavour formation during the manufacture of cheddar and, by analogy, other similar varieties of cheese. The importance of phenotypic discriminative monitoring of the dominant lactobacilli present during ripening to confirm adjunct establishment and population complexity was highlighted as was the requirement to establish the metabolic attributes of the non-starter population in uninoculated control cheeses in comparative trials.     

Impact of two starter cultures on proteolysis in Kashkaval cheese

Summary Kashkaval cheese is produced with a traditional yogurt starter culture S. thermophilus 13a + Lb. delbrueckii ssp. bulgaricus 2–11. The present paper has investigated the contribution of a selected starter culture (Lactococcus lactis ssp. lactis C11 + Streptococcus thermophilus P23 + Lactobacillus casei ssp. casei RP5) for proteolysis of Kashkaval (A) in comparison with a traditional yogurt culture (B) and soluble nitrogen (at pH 4.6 and in 12% TCA) and free amino acids as indicators of Kashkaval ripeness. After the microflora was isolated and differentiated, the largest count of mesophilic cocci (69%) and lower counts of thermophilic cocci (19.8%) and mesophilic lactobacilli (11.2%) were found in cheddared curd A, whereas in cheddared curd B thermophilic cocci (69%) prevailed over thermophilic lactobacilli (30%). Hot-brining of curd at 72 °C for 2 min destroyed 91 and 84.5%, respectively, of the starter microflora A and B and two batches of Kashkaval started ripening with 3.9 × 10⁸ c.f.u. g⁻¹ and 3.4 × 10⁸ c.f.u. g⁻¹. The ripening of Kashkaval cheese occurred mainly under the influence of the starter microflora. Ripening of Kashkaval A occurred with dominating presence of Lactobacillus casei ssp. casei RP5 from 64% at day 30 to 95.2% at day 90. In Kashkaval B the participation of Lactobacillus delbrueckii ssp. bulgaricus 2–11 increased from 36.3 to 39.9%, however it always remained relatively smaller (1.5–2.3-fold) than that of thermophilic cocci. Great activity of transformation of pH 4.6-soluble nitrogen into TCA-soluble nitrogen in Kashkaval A was registered. The high proteolytic activity of starter culture A shortened the ripening process by 30 days (30-day-old Kashkaval A had 17.3% level of proteolysis, 29.9% depth of proteolysis). Kashkaval B reached satisfactory ripeness properties when it was 60 days old (20.2% level of proteolysis, 24.9% depth of proteolysis). The high activity of amino acid release mainly by Lactobacillus casei ssp. casei RP5 and to a lesser extent by the cocci caused accumulation of a considerable amount of free amino acids in ripe Kashkaval A (421.9 mg (100 g)⁻¹ for 30-day-old Kashkaval). In ripe 30-day-old Kashkaval A the concentration of free amino acids was three times higher than in Kashkaval B, with domination of lysine, leucine, phenylalanine, valine and threonine.     

Lactobacillus casei metabolic potential to utilize citrate as an energy source in ripening cheese: a bioinformatics approach

AIMS: To identify potential pathways for citrate catabolism by Lactobacillus casei under conditions similar to ripening cheese. METHODS AND RESULTS: A putative citric acid cycle (PCAC) for Lact. casei was generated utilizing the genome sequence, and metabolic flux analyses. Although it was possible to construct a unique PCAC for Lact. casei, its full functionality was unknown. Therefore, the Lact. casei PCAC was evaluated utilizing end-product analyses of citric acid catabolism during growth in modified chemically defined media (mCDM), and Cheddar cheese extract (CCE). Results suggest that under energy source excess and limitation in mCDM this micro-organism produces mainly L-lactic acid and acetic acid, respectively. Both organic acids were produced in CCE. Additional end products include D-lactic acid, acetoin, formic acid, ethanol, and diacetyl. Production of succinic acid, malic acid, and butanendiol was not observed. CONCLUSIONS: Under conditions similar to those present in ripening cheese, citric acid is converted to acetic acid, L/D-lactic acid, acetoin, diacetyl, ethanol, and formic acid. The PCAC suggests that conversion of the citric acid-derived pyruvic acid into acetic acid, instead of lactic acid, may yield two ATPs per molecule of citric acid. Functionality of the PCAC reductive route was not observed. SIGNIFICANCE AND IMPACT OF THE STUDY: This research describes a unique PCAC for Lact. casei. Additionally, it describes the citric acid catabolism end product by this nonstarter lactic acid bacteria during growth, and under conditions similar to those present in ripening cheese. It provides insights on pathways preferably utilized to derive energy in the presence of limiting carbohydrates by this micro-organism.     

Understanding plant rooting patterns in semi‐arid systems: an integrated model analysis of climate,soil type and plant biomass

Aim A consistent set of root characteristics for herbaceous plants growing in water‐limited environments has been developed based on compilations of global root databases, but an overall analysis of why these characteristics occur is still missing. The central question in this study is whether an ecohydrological model which assumes that rooting strategies reflect maximization of transpiration can predict the variations in rooting strategies of plants in dry environments. Location Arid ecosystems across the globe. Methods A model was used to explore interactions between plant biomass, root–shoot allocation, root distribution, rainfall, soil type and water use by plants. Results Model analyses showed that the predicted shifts in rooting depth and root–shoot allocation due to changes in rainfall, soil type and plant biomass were quite similar to observed shifts. The model predicted that soil type, annual rainfall and plant biomass each had strong effects on the rooting strategies that optimize transpiration, but also that these factors have strong interactive effects. The process by which plants compete for water availability (soil evaporation or drainage) especially affected the depth distribution of roots in the soil, whereas the availability of rainfall mainly affected the optimal root–shoot allocation strategy. Main conclusions The empirically observed key patterns in rooting characteristics of herbaceous plant species in arid environments could be explained in this theoretical study by using the concept of hydrological optimality, represented here by the maximization of transpiration.     

Treatability of cheese whey for single-cell protein production in nonsterile systems: Part I. Optimal condition for lactic acid fermentation using a microaerobic sequencing batch reactor (microaerobic SBR) with immobilized Lactobacillus plantarum TISTR 2265 and microbial communities

Cheese whey contains a high organic content and causes serious problems if it is released into the environment when untreated. This study aimed to investigate the optimum condition of lactic acid production using the microaerobic sequencing batch reactor (microaerobic SBR) in a nonsterile system. The high production of lactic acid was achieved by immobilized Lactobacillus plantarum TISTR 2265 to generate an acidic pH condition below 4.5 and then to support single-cell protein (SCP) production in the second aerobic sequencing batch reactor (aerobic SBR). A hydraulic retention time (HRT) of 4 days and a whey concentration of 80% feeding gave a high lactic acid yield of 12.58 g/L, chemical oxygen demand (COD) removal of 62.38%, and lactose utilization of 61.54%. The microbial communities in the nonsterile system were dominated by members of lactic acid bacteria, and it was shown that the inoculum remained in the system up to 330 days.     

Defining patch contribution in source-sink metapopulations: the importance of including dispersal and its relevance to marine systems

In metapopulations, individual patch contribution (source or sink) is typically calculated as a patch growth rate (the intrinsic lambda, _I) dependent only upon local demographics. We demonstrate that when dispersal is explicitly included in the model, the growth rates for all patches calculated in an analogous manner (the observed lambda, _O) equilibrate to the overall metapopulation growth rate and thus no longer serve as a useful reflection of the demographic and dispersive characteristics of a given patch. In these situations we suggest an alternative method of estimating patch contribution (the contribution lambda, _C) in which a patch is decremented for losses that occur within it and credited for gains that occur anywhere in the metapopulation because of it. We compare values of _I, _O, and _C for individual patches in discrete-time density-independent metapopulation models of two organisms with very different life histories, mayflies with adult dispersal, and reef fish with larval dispersal. Results confirm that when dispersal is included only _C clearly indicates the contribution of a particular patch. _I–_C comparisons indicate that inclusion of dispersal in the mayfly model was only important if connectivity patterns were random or directional. In the reef fish model, however, results were very different when dispersal was included and there were many cases of patches being misidentified (e.g., as a source when it was really a sink) depending upon the metric used (_I or _C). Our results demonstrate the importance of including dispersal in metapopulation models when considering the contribution of individual patches.     

Application of microelectrode technique to measure pH and oxidoreduction potential gradients in gelled systems as model food

A microelectrode system is used in order to simultaneously measure pH and oxidoreduction potential (Eh) gradients developed during growth by Escherichia coli and Lactobacillus plantarum immobilized in gelled media used as model food. Unlike E. coli, L. plantarum steadily decreased medium pH independently of depth of measurement and time of incubation. Both bacteria brought about the creation of an Eh gradient throughout the gelled medium. This gradient was much more important for E. coli (700 mV) than for L. plantarum (80 mV) but more transitory.     

Role of α‐tocopherol and Lactobacillus plantarum in the alleviation of mercuric chloride‐induced testicular atrophy in rat's model: Implication of molecular mechanisms

The present work was aimed to evaluate the protective effects of alpha‐tocopherol (α‐toco) and/or Lactobacillus plantarum (LCB) against testicular atrophy induced by mercuric chloride (MCH). Rats were injected with 5 mg/kg MCH for 5 days consecutively, then treated with 100 mg/kg α‐toco and 6 × 10¹⁰ CFU 1.8701/kg LCB alone or together for 3 weeks. The MCH elevated serum TNF‐α, IL‐ 6, caspase‐3, and testicular malondialdehyde. However, serum testosterone, dehydroepiandrosterone, testicular messenger RNA of a steroidogenic acute regulatory protein, 17‐β‐hydroxysteroid dehydrogenase, 3β‐hydroxysteroid dehydrogenase, glutathione level, and superoxide dismutase activity were decreased. Protein expression of Nrf2 was downregulated whereas that of Bax and DNA fragmentation was upregulated in the testicular tissues. Treatment with α‐toco and LCB ameliorated the deviated biochemical parameters and improved tissue injury. It was concluded that the combination of LCB and α‐toco achieved promising results in the amelioration of MCH‐induced testicular atrophy. Nrf2, Bax expressions, and DNA fragmentation are involved in the testicular atrophy induced by MCH.     

Lack of biofilm contribution to bacterial colonisation in an experimental model of foreign body infection by Staphylococcus aureus and Staphylococcus epidermidis

The contribution of in vivo biofilm-forming potential of Staphylococcus aureus and Staphylococcus epidermidis was studied in an experimental model of foreign body infections. Increasing inocula (from 10(2) to 10(7) organisms) of ica-positive strains of S. aureus and S. epidermidis and their ica-negative isogenic mutants (the ica locus codes for a major polysaccharide component of biofilm) were injected into subcutaneously implanted tissue cages in guinea pigs. Surprisingly, bacterial counts and time-course of tissue cage infection by ica-positive strains of S. aureus or S. epidermidis were equivalent to those of their respective ica-negative mutants, in the locally infected fluids and on tissue-cage-inserted plastic coverslips.     

Responses of two species of heathland rodents to habitat manipulation: Vegetation density thresholds and the habitat accommodation model

The abundance of two native rodent species, Rattus lutreolus and Pseudomys gracilicaudatus, has been shown to correlate with vegetation density in coastal wet heath. Fox's habitat accommodation model relates relative abundances of such small mammal species to heathland vegetation regeneration following disturbance. Implicit in the model is recognition that it is successional changes in vegetation, not time per se, that drives the responses of small mammal species along a regeneration axis. Using a brush‐cutter we deliberately removed approximately 85% of vegetation around trapping stations and recorded significant reductions in the abundance of both P. gracilicaudatus (an earlier‐stage colonizing species) and R. lutreolus (a late seral‐stage species). A significant decrease in the abundance of only the latter had been demonstrated previously when 60–70% of the vegetation had been removed. Following the brush‐cutting both species re‐entered the mammalian secondary succession at different times, first P. gracilicaudatus followed by R. lutreolus after the vegetation cover thresholds of each species had been reached. The impact of this habitat manipulation experiment was to produce a retrogression of the small mammal succession, experimentally demonstrating causality between changes in vegetation density and subsequent small mammal habitat use.     

The contribution of marine aggregate‐associated bacteria to the accumulation of pathogenic bacteria in oysters: an agent‐based model

Bivalves process large volumes of water, leading to their accumulation of bacteria, including potential human pathogens (e.g., vibrios). These bacteria are captured at low efficiencies when freely suspended in the water column, but they also attach to marine aggregates, which are captured with near 100% efficiency. For this reason, and because they are often enriched with heterotrophic bacteria, marine aggregates have been hypothesized to function as important transporters of bacteria into bivalves. The relative contribution of aggregates and unattached bacteria to the accumulation of these cells, however, is unknown. We developed an agent‐based model to simulate accumulation of vibrio‐type bacteria in oysters. Simulations were conducted over a realistic range of concentrations of bacteria and aggregates and incorporated the dependence of pseudofeces production on particulate matter. The model shows that the contribution of aggregate‐attached bacteria depends strongly on the unattached bacteria, which form the colonization pool for aggregates and are directly captured by the simulated oysters. The concentration of aggregates is also important, but its effect depends on the concentration of unattached bacteria. At high bacterial concentrations, aggregates contribute the majority of bacteria in the oysters. At low concentrations of unattached bacteria, aggregates have a neutral or even a slightly negative effect on bacterial accumulation. These results provide the first evidence suggesting that the concentration of aggregates could influence uptake of pathogenic bacteria in bivalves and show that the tendency of a bacterial species to remain attached to aggregates is a key factor for understanding species‐specific accumulation.     

The model of Monod, Wyman, and Changeux generates two sets of parameters when applied to oxygen binding in hemoglobin

The model of Monod, Wyman and Changeux is applied to binding phenomena where the Mass Law and its expansion are employed. In this communication the model of Monod, Wyman and Changeux (MWC) is applied to analyze the oxygen binding reaction in hemoglobin. The symmetrical structure of the MWC model with its three parameters is such that two sets of these parameters, rather than one, fit experimental data for the binding of oxygen to hemoglobin.     

Dopamine transporter relation to levodopa-derived synaptic dopamine in a rat model of Parkinson's: an in vivo imaging study

Studies showed that the dopamine (DA) transporter (DAT) modulates changes in levodopa-derived synaptic dopamine levels (Δ(DA)) in Parkinson's disease (PD). Here we evaluate the relationship between DAT and Δ(DA) in the 6-hydroxydopamine model of Parkinson's disease to investigate these mechanisms as a function of dopaminergic denervation and in relation to other denervation-induced regulatory changes. 27 rats with a unilateral 6-hydroxydopamine lesion (denervation ∼20–97%) were imaged with ¹¹C-dihydrotetrabenazine (VMAT2 marker), ¹¹C-methylphenidate (DAT marker) and ¹¹C-raclopride (D2-type receptor marker). For denervation <75%Δ(DA) was significantly correlated with a combination of relatively preserved terminal density and lower DAT. For denervation <90%, Δ(DA) was significantly negatively correlated with DAT with a weaker dependence on VMAT2. For the entire data set, no dependence on pre-synaptic markers was observed; Δ(DA) was significantly positively correlated with ¹¹C-raclopride binding-derived estimates of DA loss. These findings parallel observations in humans, and show that (i) regulatory changes attempt to normalize synaptic DA levels (ii) a lesion-induced functional dependence of Δ(DA) on DAT occurs up to ∼ 90% denervation (iii) for denervation < 75% relative lower DAT levels may relate to effective compensation; for higher denervation, lower DAT levels likely contribute to oscillations in synaptic DA associated with dyskinesias.     

Progress in tissue culture,genetic transformation and applications of biotechnology to trees: an overview

Trees are an integral part of human life, and a vital component of biodiversity. Forest trees in particular are renewable sources of food, fodder, fuel wood, timber and other valuable non-timber products. Due to the rapid growth of population and the human desire to progress, there has been a tremendous reduction in forest cover from the earths surface. To maintain and sustain forest vegetation, conventional approaches have been exploited in the past for propagation and improvement. However, such efforts are confronted with several inherent bottlenecks. Biotechnological interventions for in vitro regeneration, mass micropropagation and gene transfer methods in forest tree species have been practised with success, especially in the last decade. Against the background of the limitations of long juvenile phases and life span, development of plant regeneration protocols and genetic engineering of tree species are gaining importance. Genetic engineering assumes additional significance, because of the possibility of introducing a desired gene in a single step for precision breeding of forest trees. There are no comprehensive and detailed reviews available combining research developments with major emphases on tissue culture and basic genetic transformation in tree species. The present communication attempts to overview the progress in tissue culture, genetic transformation and biotechnological applications in the last decade and future implications.     

1961-2010年内蒙古草原植被分布和生产力变化——基于MaxEnt模型和综合模型的模拟分析

定量评估气候变化对内蒙古草原植被分布及其净第一性生产力的影响有助于理解干旱区域生态系统结构和功能对气候变化的响应。基于最大熵模型（MaxEnt）评价了气候因子的重要性,进而模拟了1961-2010年内蒙古草原植被的地理分布,同时应用综合模型模拟了净第一性生产力变化。研究表明,湿润指数（MI）、年降水量（P）、最暖月平均温度（T_w）和最冷月平均温度（T_c）是决定草原植被分布的主导气候因子。1961-2010年内蒙古草甸草原、典型草原和荒漠草原分布面积分别减少了5%、1%和62%,草原面积整体减少了11%,预示着草原向着荒漠化的方向发展。降水是决定内蒙古草原净第一性生产力变化的最重要因素。     

The shapes of neutral gene genealogies in two species: probabilities of monophyly,paraphyly, and polyphyly in a coalescent model

Abstract.— The genealogies of samples of orthologous regions from multiple species can be classified by their shapes. Using a neutral coalescent model of two species, I give exact probabilities of each of four possible genealogical shapes: reciprocal monophyly, two types of paraphyly, and polyphyly. After the divergence that forms two species, each of which has population size N , polyphyly is the most likely genealogical shape for the lineages of the two species. At ∼ 1.300 N generations after divergence, paraphyly becomes most likely, and reciprocal monophyly becomes most likely at ∼1.665 N generations. For a given species, the time at which 99% of its loci acquire monophyletic genealogies is ∼5.298 N generations, assuming all loci in its sister species are monophyletic. The probability that all lineages of two species are reciprocally monophyletic given that a sample from the two species has a reciprocally monophyletic genealogy increases rapidly with sample size, as does the probability that the most recent common ancestor (MRCA) for a sample is also the MRCA for all lineages from the two species. The results have potential applications for the testing of evolutionary hypotheses.     

Prediction of desiccation sensitivity in seeds of woody species: a probabilistic model based on two seed traits and 104 species

BACKGROUND AND AIMS: Seed desiccation sensitivity limits the ex situ conservation of up to 47 % of plant species, dependent on habitat. Whilst desirable, empirically determining desiccation tolerance levels in seeds of all species is unrealistic. A probabilistic model for the rapid identification of woody species at high risk of displaying seed desiccation sensitivity is presented. METHODS: The model was developed using binary logistic regression on seed trait data [seed mass, moisture content, seed coat ratio (SCR) and rainfall in the month of seed dispersal] for 104 species from 37 families from a semi-deciduous tropical forest in Panamá. KEY RESULTS: For the Panamanian species, only seed mass and SCR were significantly related to the response to desiccation, with the desiccation-sensitive seeds being large and having a relatively low SCR (i.e. thin 'seed' coats). Application of this model to a further 38 species, of known seed storage behaviour, from two additional continents and differing vegetation types (dryland Africa and temperate Europe) correctly predicted the response to desiccation in all cases, and resolved conflicting published data for two species (Acer pseudoplatanus and Azadirachta indica). CONCLUSIONS: This model may have application as a decision-making tool in the handling of species of unknown seed storage behaviour in species from three disparate habitats.     

Establishment and comparative analyses of different culture conditions of primary hepatocytes from Nile tilapia (Oreochromis niloticus) as a model to study stress induction in vitro

Summary We established an in vitro hepatocyte primary culture system from Oreochromis niloticus, a tropical fish species of great economical importance, and evaluated its ability to express albumin, a liver-specific protein, consistently for a period of 3 wk. Serum requirements for fish hepatocyte cultures were assessed. A one-step in situ perfusion of tilapia liver retrogradely followed by collagenase liver dissociation and subsequent washing produced nearly 90% homogenous viable hepatocytes, as shown by trypan blue exclusion test. Mixed primary monolayer and aggregate hepatocyte cultures achieved by 10% fetal calf serum medium supplements expressed consistent levels of albumin. The results of light and electron microscopy showed that the hepatocytes did not significantly proliferate (P<0.05) but remained viable for at least 3 wk. The results of this study show that in vitro cultures of mixed primary hepatocyte monolayers and aggregates established from Nile tilapia may be useful models for studying transient cellular stress induction.     

Independence and interdependence in collective decision making: an agent-based model of nest-site choice by honeybee swarms

Condorcet''s jury theorem shows that when the members of a group have noisy but independent information about what is best for the group as a whole, majority decisions tend to outperform dictatorial ones. When voting is supplemented by communication, however, the resulting interdependencies between decision makers can strengthen or undermine this effect: they can facilitate information pooling, but also amplify errors. We consider an intriguing non-human case of independent information pooling combined with communication: the case of nest-site choice by honeybee (Apis mellifera) swarms. It is empirically well documented that when there are different nest sites that vary in quality, the bees usually choose the best one. We develop a new agent-based model of the bees'' decision process and show that its remarkable reliability stems from a particular interplay of independence and interdependence between the bees.