Application of Lactobacillus johnsonii expressing phage endolysin for control of Clostridium perfringens.

Clostridium perfringens is frequently found in food and the environment and produces potent toxins that have a negative impact on both human and animal health and particularly on the poultry industry. Lactobacillus johnsonii FI9785, isolated from the chicken gastrointestinal tract, has been demonstrated to exclude Cl. perfringens in poultry. We have investigated the interaction of wild-type Lact. johnsonii FI9785 or an engineered strain expressing a cell wall-hydrolysing endolysin with Cl. perfringens in vitro, using a batch culture designed to simulate human gastrointestinal tract conditions. Co-culture experiments indicated that acid production by Lact. johnsonii is important in pathogen control. The co-culture of the endolysin-secreting Lact. johnsonii with Cl. perfringens showed that the engineered strain had the potential to control the pathogen, but the ability to reduce Cl. perfringens numbers was not consistent. Results obtained indicate that survival of high numbers of Lact. johnsonii will be essential for effective pathogen control. Significance and impact of the study: The bacterium Lactobacillus johnsonii FI9785 reduces numbers of the pathogen Clostridium perfringens in vitro. Biocontrol was improved by engineering the strain to produce and export a cell wall-hydrolysing endolysin, but good survival of the producer strain is essential. The production of bacteriophage endolysins by commensal bacteria has the potential to improve competitive exclusion of pathogens in the gastrointestinal tract.

Does the light influence astaxanthin production in Xanthophyllomyces dendrorhous?

Astaxanthin (C40H52O4) is an important natural pigment that has considerable promising applications in human health. Until now, many efforts were made aimed to develop economically sustainable bioprocesses alternative to the chemical synthesis, to satisfy the increasing demand of this ketocarotenoid from feed, food and cosmetic industries. The extraction of natural astaxanthin from the yeast Xanthophyllomyces dendrorhous till now seems to be rather expensive if compared with chemically synthesized astaxanthin. In this article, astaxanthin production by Xanthophyllomyces dendrorhous under two different conditions was studied: a first effort was made using a conventional reactor while a second using an enlightened one. This research was aimed also to optimise astaxanthin production by testing the influence of the light and of some nutrient sources. From fermentation tests, an astaxanthin yield ranging about 970 µg g(-1) was obtained after fed batch cultivation in the conventional reactor. In the enlightened reactor lower values, about 930 µg g(-1), were found.

Production of feruloyl esterases and xylanases by Talaromyces stipitatus and Humicola grisea var. thermoidea on industrial food processing by-products.

Feruloyl esterase (FAE) and xylanase activities were detected in culture supernatants from Humicola grisea var. thermoidea and Talaromyces stipitatus grown on brewers' spent grain (BSG) and wheat bran (WB), two agro-industrial by-products. Maximum activities were detected from cultures of H. grisea grown at 150 rpm, with 16.9 U/ml and 9.1 U/ml of xylanase activity on BSG and WB, respectively. Maximum FAE activity was 0.47 U/ml and 0.33 U/ml on BSG and WB, respectively. Analysis of residual cell wall material after microbial growth shows the preferential solubilisation of arabinoxylan and cellulose, two main polysaccharides present in BSG and WB. The production of low-cost cell-wall-deconstructing enzymes on agro-industrial by-products could lead to the production of low-cost enzymes for use in the valorisation of food processing wastes.

Synergy between xylanases from glycoside hydrolase family 10 and family 11 and a feruloyl esterase in the release of phenolic acids from cereal arabinoxylan.

The bioconversion of waste residues (by-products) from cereal processing industries requires the cooperation of enzymes able to degrade xylanolytic and cellulosic material. The type A feruloyl esterase from Aspergillus niger, AnFaeA, works synergistically with (1-->4)-beta-D-xylopyranosidases (xylanases) to release monomeric and dimeric ferulic acid (FA) from cereal cell wall-derived material. The esterase was more effective with a family 11 xylanase from Trichoderma viride in releasing FA and with a family 10 xylanase from Thermoascus aurantiacus in releasing the 5,5' form of diferulic acid from arabinoxylan (AX) derived from brewers' spent grain. The converse was found for the release of the phenolic acids from wheat bran-derived AXs. This may be indicative of compositional differences in AXs in cereals.

SCP and crude pectinase production by slurry-state fermentation of lemon pulps.

Single cell protein (SCP) and crude pectinolytic enzymes production from citrus pulps is reported. SCP and enzymes were produced by slurry-state flask cultivation of Aspergillus niger and Trichoderma viride on pulps from lemon juice clarification. Production as well as crude pectinase activity was not affected by the high dry matter content of the pulps. Both the protein content in the residue and the enzyme activity in the supernatant were higher in T. viride than in A. niger culture. The crude pectinase of T. viride, whose specific activity was similar to that found for a commercial concentrated preparation, could be utilized in the same citrus processing factory as well as in other factories which use large amounts of pectinolytic crude preparations, for example to enhance depuration plant performance.

Yeast production from virgin grape marc.

An alternative utilization of virgin grape marc (VGM) to produce SCP from S. ceretisiae is reported. A simple extraction method of fresh grape marc produces a sugar-rich solution: through fed-batch fermentation, a high-value yeast biomass instead of a low-value product like ethanol can be produced. Productivity and quality of yeast are similar to these obtainable from molasses. The convenience of yeast production from VGM is briefly discussed; it appears of great interest in south Italy and generally in grape-producing countries, specially if these lack relevant sources of fermentable sugars.

Advantages and disadvantages of aggregating fluxes into synthetic and degradative fluxes when modelling metabolic pathways.

It is now widely accepted that mathematical models are needed to predict the behaviour of complex metabolic networks in the cell, in order to have a rational basis for planning metabolic engineering with biotechnological or therapeutical purposes. The great complexity of metabolic networks makes it crucial to simplify them for analysis, but without violating key principles of stoichiometry or thermodynamics. We show here, however, that models for branched complex systems are sometimes obtained that violate the stoichiometry of fluxes at branch points and as a result give unrealistic metabolite concentrations at the steady state. This problem is especially important when models are constructed with the S-system form of biochemical systems theory. However, the same violation of stoichiometry can occur in metabolic control analysis if control coefficients are assumed to be constant when trying to predict the effects of large changes. We derive the appropriate matrix equations to analyse this type of problem systematically and to assess its extent in any given model.

Mathematical models of purine metabolism in man.

Experimental and clinical data on purine metabolism are collated and analyzed with three mathematical models. The first model is the result of an attempt to construct a traditional kinetic model based on Michaelis-Menten rate laws. This attempt is only partially successful, since kinetic information, while extensive, is not complete, and since qualitative information is difficult to incorporate into this type of model. The data gaps necessitate the complementation of the Michaelis-Menten model with other functional forms that can incorporate different types of data. The most convenient and established representations for this purpose are rate laws formulated as power-law functions, and these are used to construct a Complemented Michaelis-Menten (CMM) model. The other two models are pure power-law-representations, one in the form of a Generalized Mass Action (GMA) system, and the other one in the form of an S-system. The first part of the paper contains a compendium of experimental data necessary for any model of purine metabolism. This is followed by the formulation of the three models and a comparative analysis. For physiological and moderately pathological perturbations in metabolites or enzymes, the results of the three models are very similar and consistent with clinical findings. This is an encouraging result since the three models have different structures and data requirements and are based on different mathematical assumptions. Significant enzyme deficiencies are not so well modeled by the S-system model. The CMM model captures the dynamics better, but judging by comparisons with clinical observations, the best model in this case is the GMA model. The model results are discussed in some detail, along with advantages and disadvantages of each modeling strategy.

Analysis of abnormalities in purine metabolism leading to gout and to neurological dysfunctions in man.

A modelling approach is used to analyse diseases associated with purine metabolism in man. The specific focus is on deficiencies in two enzymes, hypoxanthine:guanine phosphoribosyltransferase and adenylosuccinate lyase. These deficiencies can lead to a number of symptoms, including neurological dysfunctions and mental retardation. Although the biochemical mechanisms of dysfunctions associated with adenylosuccinate lyase deficiency are not completely understood, there is at least general agreement in the literature about possible causes. Simulations with our model confirm that accumulation of the two substrates of the enzyme can lead to significant biochemical imbalance. In hypoxanthine:guanine phosphoribosyltransferase deficiency the biochemical mechanisms associated with neurological dysfunctions are less clear. Model analyses support some old hypotheses but also suggest new indicators for possible causes of neurological dysfunctions associated with this deficiency. Hypoxanthine:guanine phosphoribosyltransferase deficiency is known to cause hyperuricaemia and gout. We compare the relative importance of this deficiency with other known causes of gout in humans. The analysis suggests that defects in the excretion of uric acid are more consequential than defects in uric acid synthesis such as hypoxanthine:guanine phosphoribosyltransferase deficiency.

Validation and steady-state analysis of a power-law model of purine metabolism in man.

The paper introduces a model of human purine metabolism in situ. Chosen from among several alternative system descriptions, the model is formulated as a Generalized Mass Action system within Biochemical Systems Theory and validated with analyses of steady-state and dynamic characteristics. Eigenvalue and sensitivity analyses indicate that the model has a stable and robust steady-state. The model quite accurately reproduces numerous biochemical and clinical observations in healthy subjects as well as in patients with disorders of purine metabolism. These results suggest that the model can be used to assess biochemical and clinical aspects of human purine metabolism. It provides a means of exploring effects of enzyme deficiencies and is a potential tool for identifying steps of the pathway that could be the target of therapeutical intervention. Numerous quantitative comparisons with data are given. The model can be used for biomathematical exploration of relationships between enzymic deficiencies and clinically manifested diseases.

Comparative characterization of the fermentation pathway of Saccharomyces cerevisiae using biochemical systems theory and metabolic control analysis: model definition and nomenclature.

Mathematical tools that involve the determination of systemic responses to small changes in metabolites or enzymes have demonstrated their utility for analyzing metabolic pathways. The different methodologies based on these ideas allow for modeling and analyzing biochemical pathways focusing on the coordinate behavior of the whole system. However, one must become familiar with the difference in nomenclature and methodology to relate the models and results obtained by applying these techniques and to appreciate their potential for answering fundamental questions about biochemical systems. In the following three papers we show how this can be facilitated by comparing the nomenclature, methodology, and results of the two leading techniques in this area, metabolic control analysis and biochemical systems theory, using a model of the fermentation pathway in Saccharomyces cerevisiae as a reference system. In the present paper we review the nomenclature, technical concepts, and related experimental measurements while creating a practical dictionary for the reference system that makes the relatedness of the two approaches more apparent. In the second paper, subtitled Steady-State Analysis, we show that both approaches give the same picture for many systemic responses of the reference system. In the third paper of this series, subtitled Model Validation and Dynamic Behavior, we show that the quality of the model can be assessed by studying the sensitivity to changes in the system parameters. We hope to illustrate the usefulness of these tools in providing an interpretation of the experimental measurements in a specific metabolic pathway.

Comparative characterization of the fermentation pathway of Saccharomyces cerevisiae using biochemical systems theory and metabolic control analysis: steady-state analysis.

In the preceding paper in this issue, we have shown that metabolic control analysis and biochemical systems theory use the same experimental information to describe a metabolic system. In this paper, we analyze the steady-state properties of this pathway by applying both methods. Our results show the correspondence of the steady-state characterizations and illustrate the relationships between the different nomenclatures used. With both approaches, we identify metabolite pools that are strongly influenced by changes in enzyme concentration when cells are immobilized at pH 5.5. In the final paper of this series, which follows, we discuss the need to assess the quality of a model and the potential difficulties that may arise if the steady-state characterization is accepted without testing its quality. We then validate the different models using parameter sensitivity concepts.

Comparative characterization of the fermentation pathway of Saccharomyces cerevisiae using biochemical systems theory and metabolic control analysis: model validation and dynamic behavior.

In the first two papers of this series (immediately preceding, this issue), we characterized the steady-state properties of a model of a fermentation pathway in Saccharomyces cerevisiae in four experimental conditions. In each of these conditions, the pictures obtained by metabolic control analysis and biochemical systems theory were coincident, which illustrates the relatedness of the two approaches. In this paper we analyze the quality of this description by means of the tools available within biochemical systems theory, and we show that in some of the experimental conditions studied the system is poorly characterized. The most critical condition corresponds to the immobilization of the cells at pH 5.5, in which the kinetic characterization appears to be inaccurate. Furthermore, sensitivity analysis and the study of the local steady-state stability identify the most critical parameters. The results of these analyses are confirmed by the predictions of the dynamic response of the model using its S-system representation. This illustrates the utility of these tools and warns against using the steady-state characterization without testing its validity.

Biological effects of hesperidin, a Citrus flavonoid. (note II): hypolipidemic activity on experimental hypercholesterolemia in rat.

Hesperidin, the most important flavanone of Citrus sp., significantly increases HDL and lowers cholesterol, LDL, total lipid and triglyceride plasma levels in normolipidemic rats and in rats with diet- and triton-induced hyperlipidemia.

[A case of sepsis of the urinary tract caused by Candida albicans]. / Un caso di sepsi delle vie urinarie sostenuto da "Candida albicans"

The AA. describe a case of secondary sepsis of the urinary tract by Candida albicans. The subject, after an operation of right inguinal herniotomy, was treated prophylactically with Ceporin. Three days after the operation he was catheterized and Bactrin was added to Ceporin. It was possible to isolate from urine a strain of Candida. Morphological, cultural, biochemical features, as well as fermentation and assimilation tests, and the biological assay of pathogenicity on the rabbit, allowed the strain of the yeast to be identified as Candida albicans. Moreover, an extract obtained from the yeast, tested by Wadsworth's micromethod against the serum of the patient, showed two lines of precipitation. The patient recovered after treatment with antimycotics.