An H2-T MHC class Ib molecule presents Listeria monocytogenes-derived antigen to immune CD8+ cytotoxic T cells

Mouse spleen T cells can adoptively transfer immunity to Listeria monocytogenes; this activity was markedly enhanced by stimulation with Con A in vitro before transfer. The enhanced and prolonged protection against L. monocytogenes in vivo was correlated with enhanced lysis in vitro of target cells infected with strains of L. monocytogenes that produce listeriolysin O (LLO). One of the targets of such cytotoxic cells from BALB/c (H2d) mice was a peptide that corresponded to amino acids 91 to 99 (p91-99) of the LLO molecule, which satisfies the binding motif of H2-Kd. Listeria-immune CD3+CD8+, but not CD3+CD8-, cells could also lyse H-2-incompatible, infected target cells. Immune cells from C57BL/6 (H2b) mice lysed allogeneic H-2d target cells infected with L. monocytogenes or a Bacillus subtilis transformant that secretes LLO, but did not lyse targets pulsed with p91-99. This H2-unrestricted cytolysis was therefore directed at a fragment of the LLO molecule other than p91-99. Listeria-infected bone marrow macrophages from congenic and recombinant strains of mice were lysed only when they shared the H2-T region or were Qa1-compatible with the immune cytotoxic cells; sharing of the H2-D, Q, or M region was insufficient. Thus, the immune response to L. monocytogenes included cytolytic CD8+ cells that recognized endogenously processed Listeria-derived Ags in the context of the class Ia H2-K molecule, as well as a class Ib H2-T molecule.     

Kinetics of the thermal inactivation of the Lactococcus lactis bacteriophage P008

The thermal resistance of the lactococcal bacteriophage P008 was investigated between 55 and 80 degrees C. Inactivation kinetics revealed an order of reaction above 1 and could be determined by a non-1st-order regression model. Phage inactivation was influenced by the medium (milk and Ca-M17-broth). Within the investigated temperature range, milk had a protective effect on phage P008. This was reflected in the rate constant and in the activation energy. Thermal phage inactivation studies reported in literature were re-analysed using non-1st-order regression. The obtained kinetic parameters showed that phage P008 belongs to the most heat resistant lactococcal phages investigated so far.     

Developmental competence in vivo and in vitro of in vitro-matured equine oocytes fertilized by intracytoplasmic sperm injection with fresh or frozen-thawed spermatozoa

This study was undertaken to evaluate the development of equine oocytes in vitro and in vivo after intracytoplasmic sperm injection (ICSI) with either fresh or frozen-thawed spermatozoa, without the use of additional activation treatments. Oocytes were collected from ovaries obtained from an abattoir and oocytes classified as having expanded cumulus cells were matured in M199 with 10% fetal bovine serum and 5 microU FSH ml(-1). After 24-26 h of in vitro maturation, oocytes with a first polar body were selected for manipulation. Fresh ejaculated stallion spermatozoa were used for the experiment after swim-up for 20 min in sperm-Tyrode's albumen lactate pyruvate. Frozen-thawed spermatozoa from the same stallion were treated in a similar way. Spermatozoa were immobilized and injected into the oocytes using a Piezo drill. Presumptive zygotes were cultured in G1.2 medium for 20 or 96 h after the injection was administered, or were transferred to the oviducts of recipient mares and recovered 96 h later. In addition, bovine oocytes with first polar bodies were injected with the two types of stallion spermatozoa and fixed 20 h after injection to examine pronuclear formation. Fertilization rate (pronucleus formation and cleavage) at 20 h after injection of spermatozoa was not significantly different between fresh and frozen-thawed sperm groups in either equine or bovine oocytes. Pronucleus formation after injection of spermatozoa into bovine oocytes was significantly higher than that for equine oocytes (P < 0.05). There were no significant differences in cleavage rate or average number of nuclei at 96 h between equine oocytes injected with fresh or frozen-thawed spermatozoa. However, embryos developed in vivo for 96 h had a significantly higher number of nuclei in both sperm treatments compared with those cultured in vitro. These results indicate that good activation rates may be obtained after injection of either fresh or frozen-thawed equine spermatozoa without additional activation treatment. Injection of frozen-thawed equine spermatozoa results in similar embryo development to that obtained with fresh equine spermatozoa. In vitro culture of equine zygotes in G1.2 medium results in a similar cleavage rate but reduced number of cells compared with in vivo culture within the oviduct. Bovine oocytes may be useful as models for assessing sperm function in horses.     

Effect of the fat globule sizes on the meltdown of ice cream

The meltdown of ice cream is influenced by its composition and additives and by fat globule size. The objective of this study was to examine the effect of fat globule size and fat agglomerate size on the meltdown stability of ice cream. Therefore, an ice cream mix (10% milk fat) was homogenized at pressures ranging from 0 to 30 MPa in single-stage, double-stage, and selective homogenization processes. The ice cream, produced on a continuous ice cream freezer, was characterized by an optimized meltdown test while, in addition, the fat globule sizes and the free fat content were determined in the mix and the molten ice cream. The meltdown was dependent on the fat agglomerate sizes in the unfrozen serum phase. Agglomerates smaller than a critical diameter led to significantly higher meltdown rates. Homogenization pressures of at least 10 MPa were sufficient to produce a stable ice cream. Furthermore, proof was provided that double-stage homogenization is not necessary for fat contents up to 10% and that selective homogenization is possible to produce stable ice creams. Based on these results a model was deduced describing the stabilizing mechanisms during the meltdown process.     

Determination of cleaning end of dairy protein fouling using an online system combining ultrasonic and classification methods

Fouling and cleaning of heat exchangers in food industry are severe and costly issues and of high importance. In this study, a planar heat exchanger was constructed to produce and clean milk protein fouling similar to industry. Using a combination of an ultrasonic measuring method and classification machines cleaning should be monitored online to adapt cleaning time. After reproducible fouling deposit was built, cleaning started which was monitored using an ultrasonic measuring unit. The measured ultrasonic signal was analyzed for seven acoustic features and fed together with temperature and mass flow rate (both measured) into a classification method for decision of fouling presence or absence. For classification, artificial neural network (ANN) and support vector machine (SVM) was applied displaying detection accuracies of more than 80 % (ANN) and 94 % (SVM), respectively. Besides, the slope change of the seven acoustic features was monitored with time resulting in a cleaning time of at least 21?±?4 min. The cleaning time determined by the new sensor system is comparable with previously determined cleaning times for this setup. This study demonstrated that ultrasound based sensor systems offer a new tool to determine presence or absence of fouling and to monitor cleaning processes in the food industry with high accuracy.     

Initial Droplet Size Impacts pH‐Induced Structural Changes in Phase‐Separated Polymer Dispersions

The effect of pH change on the morphology of whey protein isolate (WPI)–pectin dispersions obtained from phase‐separated systems after mild shear was studied. The purpose of this study was to examine the impact of mixing speed on the initial particle size of biopolymer complexes and their structure morphology after sequentially changing the pH. Therefore, solutions of WPI and pectin were combined at pH 6.1, allowed to phase separate and were then mildly homogenized at 50, 100, and 150 rpm, respectively, to form a dispersion containing differently sized WPI droplets in a surrounding pectin‐rich phase. Each dispersion was then subjected to a pH change, such as 6.1 to 5.2 and 3.2, by slowly adding hydrochloric acid. The systems morphology, size, appearance, rheology, and storage stability was then characterized by optical microscopy, static light scattering, visual inspections, and steady shear rheometry to gain insights into the structural rearrangements. Results indicated substantial changes in the structure of the dispersion when the pH was changed. Formation of core‐shell structures from the WPI droplets was observed at an intermediate pH. There, initial droplet size was found to affect structures formed, that is, core‐shell type particles would only form if droplets were large (>1.5 μm) prior to pH change. Insights gained may be of importance to food manufacturers intending to create new structures from mixtures of proteins and carbohydrates.     

Genetic parameters for health traits and their relationship to different persistency traits in German Holstein dairy cattle

Data from 3,200 Holstein cows from 3 commercial dairy farms in Germany were used to estimate heritabilities and breeding values for liability to udder diseases (UD), fertility diseases (FD), metabolic diseases (MD), and claw and leg diseases (CLD) using single-trait threshold sire models. A total of 92,722 medical treatments recorded from 1998 to 2003 were included in the analysis. Approximate genetic correlations between persistency of milk yield, fat yield, protein yield, and persistency of milk energy yield and liability to the health traits were calculated based on correlations between EBV. Posterior means of heritability of liability ranged from 0.05 to 0.08 for UD, from 0.04 to 0.07 for FD, from 0.08 to 0.12 for MD, and from 0.04 to 0.07 for CLD. Approximate genetic correlations of the disease traits with the persistency traits were favorable, except for MD in all lactations, which were unfavorable, and UD, which were around zero. Highest correlations in the range of 0.13 to 0.46 were found between the different persistency traits and CLD.     

Production of horse foals via direct injection of roscovitine-treated donor cells and activation by injection of sperm extract

We evaluated the effects of different donor cell treatments and activation methods on production of blastocysts after equine nuclear transfer. Nuclear transfer was performed by direct injection of donor cells, using a piezo drill, and standard activation was by injection of sperm factor followed by culture with 6-dimethylaminopurine. There was no difference in blastocyst development between embryos produced with roscovitine-treated or confluent donor cells (3.6% for either treatment). Addition of injection of roscovitine or culture with cycloheximide at the time of activation did not affect blastocyst development. Overall, transfer of eight blastocysts produced using roscovitine-treated donor cells and our standard activation protocol yielded three pregnancies, of which two (25% of transferred embryos) resulted in delivery of viable foals. Flow cytometric evaluation showed that roscovitine treatment significantly increased the proportion of cells classified as small, in comparison to growth to confluence or serum deprivation, but did not significantly affect the proportion of cells in G0/G1 (2N DNA content). Transfer of one blastocyst produced using roscovitine-treated donor cells, with addition of roscovitine injection at activation, yielded one pregnancy which was lost before 114 days' gestation. Transfer to recipients of two blastocysts produced using confluent donor cells with addition of cycloheximide at activation gave no resulting pregnancies. We conclude that roscovitine treatment of donor cells yields equivalent blastocyst production after nuclear transfer to that for confluent donor cells, and that direct injection of roscovitine-treated donor cells, followed by activation using sperm extract, is compatible with efficient production of viable cloned foals.     

Effect of homogenization, microfiltration and pH on curd firmness and syneresis of curd grains

The impact of the independent variables, homogenization pressure (p₁), concentration factor of microfiltration (i) and pH on curd firmness (CF) and syneresis of curd grains was studied. Texture analysis was used to characterize CF of the rennet-induced gels. The analysis of a two-level factorial design revealed that i, p₁, pH and the interaction of i and pH had the most important influence on CF. Cutting time was therefore individually determined for each milk system using small amplitude oscillatory rheometry for generating comparable conditions for the syneresis experiments. Syneresis of curd grains with a diameter of 11 mm was followed at 35 °C close to semi-hard cheesemaking conditions. The permeate release during microfiltration was taken into consideration, allowing an evaluation of syneresis of grains made from concentrated and unconcentrated milk. It was shown that with increasing milk concentration less curd treatment time was needed to reach a certain syneresis value. Hence, total processing time in cheesemaking is decreased. Analysis of variance revealed that syneresis was affected by the individual variables. Kinetic parameters were satisfactorily estimated through regression (R²>0.98) and it was shown that milk composition and concentration due to microfiltration markedly influenced the endpoint of syneresis, RWR_max. The experiments demonstrate that microfiltration and homogenization can be combined to reach CF and syneresis comparable to untreated milk used in conventional cheesemaking. This meets one claim of the cheese industry when implementing both technologies in the manufacture process, since consistency and quality of the ripened cheese are expected to be unchanged.