Fluorescence-based characterisation of selected edible insect species: Excitation emission matrix (EEM) and parallel factor (PARAFAC) analysis.

Fluorescence spectroscopy coupled with chemometric tools is a powerful analytical method, largely used for rapid food quality and safety evaluations. However, its potential has not yet been explored in the novel food sector. In the present study, excitation emission matrices (EEMs) of 15 insect powders produced by milling insects belonging to 5 Orthoptera species (Acheta domesticus, Gryllus assimilis, Gryllus bimaculatus, Locusta migratoria, Schistocerca gregaria) from 3 different origins were investigated. Parallel factor (PARAFAC) analysis performed on the overall averaged dataset was validated for five components, highlighting the presence of five different fluorescence peaks. The presence of these peaks was confirmed on each species, suggesting that fluorescence compounds of edible insects are the same in several species. PARAFAC analysis performed on the overall averaged dataset after alternatively adding the EEM recorded from one standard compound allowed to speculate that edible insects fluorescence raises from mixtures of: tryptophan + tyrosine (PARAFAC component-1), tryptophan + tyrosine + tocopherol (PARAFAC component-2), collagen + pyridoxine + pterins (PARAFAC component-3). This study suggests that fluorescence spectroscopy may represent a powerful method for investigating composition and quality of insect-based foods.

Stability of hepatitis E virus at high hydrostatic pressure processing.

Hepatitis E virus (HEV) is the causative agent of acute and chronic hepatitis in humans. The zoonotic HEV genotype 3 is the main genotype in Europe. The foodborne transmission via consumption of meat and meat products prepared from infected pigs or wild boars is considered the major transmission route of this genotype. High hydrostatic pressure processing (HPP) is a technique, which can be used for inactivation of pathogens in food. Here, preparations of a cell culture-adapted HEV genotype 3 strain in phosphate-buffered saline (PBS) were subjected to HPP and the remaining infectivity was titrated in cell culture by counting fluorescent foci of replicating virus. A gradual decrease in infectivity was found by application of 100 to 600 MPa for 2 min. At 20 °C, infectivity reduction of 0.5 log10 at 200 MPa and 1 log10 at 400 MPa were observed. Slightly higher infectivity reduction of 1 log10 at 200 MPa and 2 log10 at 400 MPa were found by application of the pressure at 4 °C. At both temperatures, the virus was nearly completely inactivated (>3.5 log10 infectivity decrease) at 600 MPa; however, low amounts of remaining infectious virus were observed in one of three replicates in both cases. Transmission electron microscopy showed disassembled and distorted particles in the preparations treated with 600 MPa. Time-course experiments at 400 MPa showed a continuous decline of infectivity from 30 s to 10 min, leading to a 2 log10 infectivity decrease at 20 °C and to a 2.5 log10 infectivity decrease at 4 °C for a 10 min pressure application each. Predictive models for inactivation of HEV by HPP were generated on the basis of the generated data. The results show that HPP treatment can reduce HEV infectivity, which is mainly dependent on pressure height and duration of the HPP treatment. Compared to other viruses, HEV appears to be relatively stable against HPP and high pressure/long time combinations have to be applied for significant reduction of infectivity.

Non-destructive mobile monitoring of microbial contaminations on meat surfaces using porphyrin fluorescence intensities.

A non-destructive mobile system for meat quality monitoring was developed and investigated for the possible application along the whole production chain of fresh meat. Pork and lamb meat was stored at 5 °C for up to 20 days post mortem and measured with a fluorescence spectrometer. Additionally, the bacterial influence on the fluorescence signals was evaluated by different experimental procedures. Fluorescence of NADH and different porphyrins could be correlated to the growth of diverse bacteria and hence used for contamination monitoring. The increase of porphyrin fluorescence started after 9 days p.m. for pork and after 2 days p.m. for lamb meat. Based on the results, a mobile fluorescence system was built and compared with the laboratory system. The corrected function of the meat slices showed a root mean square error of 1156.97 r.u. and a mean absolute percentage error of 12.59%; for lamb the values were 470.81 r.u. and 15.55%, respectively. A mobile and non-invasive measurement system would improve the microbial security of fresh meat.

Durable fear memories require PSD-95.

Traumatic fear memories are highly durable but also dynamic, undergoing repeated reactivation and rehearsal over time. Although overly persistent fear memories underlie anxiety disorders, such as posttraumatic stress disorder, the key neural and molecular mechanisms underlying fear memory durability remain unclear. Postsynaptic density 95 (PSD-95) is a synaptic protein regulating glutamate receptor anchoring, synaptic stability and certain types of memory. Using a loss-of-function mutant mouse lacking the guanylate kinase domain of PSD-95 (PSD-95(GK)), we analyzed the contribution of PSD-95 to fear memory formation and retrieval, and sought to identify the neural basis of PSD-95-mediated memory maintenance using ex vivo immediate-early gene mapping, in vivo neuronal recordings and viral-mediated knockdown (KD) approaches. We show that PSD-95 is dispensable for the formation and expression of recent fear memories, but essential for the formation of precise and flexible fear memories and for the maintenance of memories at remote time points. The failure of PSD-95(GK) mice to retrieve remote cued fear memory was associated with hypoactivation of the infralimbic (IL) cortex (but not the anterior cingulate cortex (ACC) or prelimbic cortex), reduced IL single-unit firing and bursting, and attenuated IL gamma and theta oscillations. Adeno-associated virus-mediated PSD-95 KD in the IL, but not the ACC, was sufficient to impair recent fear extinction and remote fear memory, and remodel IL dendritic spines. Collectively, these data identify PSD-95 in the IL as a critical mechanism supporting the durability of fear memories over time. These preclinical findings have implications for developing novel approaches to treating trauma-based anxiety disorders that target the weakening of overly persistent fear memories.

Effects of different storage conditions on quality related porphyrin fluorescence signatures of pork slices.

This study evaluated the potential of fluorescence as an indicator of pork quality by determining the effects of various conditions on fluorescence signatures (excitation at 420 nm, emission at 550-750 nm). Storage of porcine musculus longissimus dorsi in PE bags led to a clear increase in porphyrin fluorescence intensity after approx. 10 d post mortem. Modified gas atmosphere (70% O(2), 30% CO(2)) inhibited the fluorescence emission of zinc protoporphyrin and protoporphyrin IX due to quenching by oxygen. Bleaching processes caused similar effects by halogen light exposure during meat storage. However, already formed signals could not be manipulated by oxygen or halogen light. Storage under vacuum reduced the quenching effects and resulted in increased fluorescence intensities. Freezing and thawing of meat samples delayed and reduced the increase in fluorescence intensity. Only minor effects could be detected at long-term frozen storage for two months. Consequently porphyrin fluorescence analysis is a potential means to indicate changes of pork quality and remaining shelf life.

Fluorimetric detection of protoporphyrins as an indicator for quality monitoring of fresh intact pork meat.

In fresh meat production fast and non-destructive quality monitoring along the distribution chain is a key aspect to guaranteeing high quality and safe products for consumption. The applicability of fluorescence spectroscopy using protoporphyrins as indicators for meat ageing was investigated. Porcine musculus longissimus dorsi (MLD) was stored in slices over 20 days at 5 and 12°C and measured every day with an excitation of 420nm and an emission range of 550-750nm. Additionally, pH, drip loss and colour were examined to assess possible correlations. The obtained spectra of the MLD showed an increase in three peaks at 592, 638 and 705nm which could be reconstructed using the spectra of standard solutions of protoporphyrin IX (PP) and zinc protoporphyrin IX (ZnPP) or magnesium protoporphyrin (MgPP), respectively. Using principal component analysis (PCA) on the fluorescence spectral data, the meat slices stored at 5°C showed differences in the fluorescence signal after the 10th day and 5th day when stored at 12°C. An interrelationship between the additional analyses and the fluorescence intensities on these relevant days could not be established. In conclusion, the increase of ZnPP fluorescence due to temperature related changes of physiological meat properties is capable of serving as a quality indicator with regards to inadequate conditioning (e.g. during transportation and/or storage) of pork meat.

Treatment of recycled carrot washing water.

Large volumes of effluents and solid waste derive from industrial fresh packing and processing of carrots. Due to high organic loads and suspended solids and due to insufficient wastewater treatment, surface waters are polluted by effluents of carrot washing, as shown for one enterprise in Germany. Over a period of four months the water quality was studied at the stages of a treatment system consisting of a grit chamber, a settling tank combined with aeration devices and a constructed wetland. Organic pollutants were measured as COD and BOD5 in different fractions of the wastewater. The daily washing of carrots produces up to 300 m3 wastewater with a mean filterable solids content of 453 mg l(-1) and a mean COD concentration of 179 mg l(-1). On average the current efficiency of total COD reduction is 52%. Due to the fine grain size and the low density of suspended solids, settling is very slow and incomplete, primarily during the winter. As much as 29 mg COD l(-1) in the effluent derive from the fraction of particles which do not settle within 18 hours. The dissolved COD fraction accounts for 30 mg l(-1) and reflects the insufficient oxygenation of the wastewater. Strategies for enhancing the removal of organic compounds can be derived from the results of this study. Spatial separation of settling and aeration processes is recommended to optimise the treatment process. Furthermore, the area of the constructed wetland should be increased to maintain its hydraulic conductivity and to guarantee a high COD reduction.

Metastable states of water and ice during pressure-supported freezing of potato tissue.

Different ice modifications were obtained during freezing processes at several pressure levels from atmospheric pressure up to 300 MPa. In the pressure range between 210 and 240 MPa, a metastable ice I modification area was observed, as the nucleation of ice I crystals in the thermodynamically stable region of ice III was reached. A significant degree of supercooling was obtained before freezing the tissue water to ice III, which has to be considered when designing pressure-supported freezing processes. The effect of supercooling phenomenon on the phase transition time is discussed using a mathematical model based on the solution of the heat transfer governing differential equations. Phase transition and freezing times for the different freezing paths experimented are compared for the processes: freezing at atmospheric pressure, pressure-assisted freezing, and pressure-shift freezing. Different metastable states of liquid water are defined according to their process-dependent stability.

Role of alpha-synuclein in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced parkinsonism in mice.

In humans, mutations in the alpha-synuclein gene or exposure to the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) produce Parkinson's disease with loss of dopaminergic neurons and depletion of nigrostriatal dopamine. alpha-Synuclein is a vertebrate-specific component of presynaptic nerve terminals that may function in modulating synaptic transmission. To test whether MPTP toxicity involves alpha-synuclein, we generated alpha-synuclein-deficient mice by homologous recombination, and analyzed the effect of deleting alpha-synuclein on MPTP toxicity using these knockout mice. In addition, we examined commercially available mice that contain a spontaneous loss of the alpha-synuclein gene. As described previously, deletion of alpha-synuclein had no significant effects on brain structure or composition. In particular, the levels of synaptic proteins were not altered, and the concentrations of dopamine, dopamine metabolites, and dopaminergic proteins were unchanged. Upon acute MPTP challenge, alpha-synuclein knockout mice were partly protected from chronic depletion of nigrostriatal dopamine when compared with littermates of the same genetic background, whereas mice carrying the spontaneous deletion of the alpha-synuclein gene exhibited no protection. Furthermore, alpha-synuclein knockout mice but not the mice with the alpha-synuclein gene deletion were slightly more sensitive to methamphetamine than littermate control mice. These results demonstrate that alpha-synuclein is not obligatorily coupled to MPTP sensitivity, but can influence MPTP toxicity on some genetic backgrounds, and illustrate the need for extensive controls in studies aimed at describing the effects of mouse knockouts on MPTP sensitivity.

Alpha-synuclein accumulation in a case of neurodegeneration with brain iron accumulation type 1 (NBIA-1, formerly Hallervorden-Spatz syndrome) with widespread cortical and brainstem-type Lewy bodies.

We studied a 27-year-old woman who died after a 6-year history of progressive dementia, dystonia, ataxia, apraxia, spasticity, choreoathetosis, visual and auditory hallucinations, and optic atrophy. Magnetic resonance imaging showed decreased intensity in the globus pallidus, substantia nigra, and dentate nuclei in T2-weighted images, supporting the clinical diagnosis of neurodegeneration with brain iron accumulation type 1 (NBIA-1; formerly known as Hallervorden-Spatz syndrome). At autopsy the brain showed mild frontotemporal atrophy and discoloration of the globus pallidus and the substantia nigra pars reticularis. Histologically, features typical of NBIA-1 were found including widespread axonal spheroids and large deposits of iron pigment in the discolored regions. Additionally, excessive numbers of Lewy bodies (LBs) were found throughout all examined brain stem and cortical regions. LBs of both types, as well as Lewy neurites in this case of NBIA-1, were strongly labeled by antibodies against alpha-synuclein. These findings give further evidence that accumulation of alpha-synuclein is generally associated with LB formation, i.e., in Parkinson's disease, dementia with Lewy bodies and NBIA-1. The case presented here is particularly notable for its high number of LBs in all areas of the cerebral cortex.

Rabphilin knock-out mice reveal that rabphilin is not required for rab3 function in regulating neurotransmitter release.

Rab3A and rab3C are GTP-binding proteins of synaptic vesicles that regulate vesicle exocytosis. Rabphilin is a candidate rab3 effector at the synapse because it binds to rab3s in a GTP-dependent manner, it is co-localized with rab3s on synaptic vesicles, and it dissociates with rab3s from the vesicles during exocytosis. Rabphilin contains two C(2) domains, which could function as Ca(2+) sensors in exocytosis and is phosphorylated as a function of stimulation. However, it is unknown what essential function, if any, rabphilin performs. One controversial question regards the respective roles of rab3s and rabphilin in localizing each other to synaptic vesicles: although rabphilin is mislocalized in rab3A knock-out mice, purified synaptic vesicles were shown to require rabphilin for binding of rab3A but not rab3A for binding of rabphilin. To test whether rabphilin is involved in localizing rab3s to synaptic vesicles and to explore the functions of rabphilin in regulating exocytosis, we have now analyzed knock-out mice for rabphilin. Mice that lack rabphilin are viable and fertile without obvious physiological impairments. In rabphilin-deficient mice, rab3A is targeted to synaptic vesicles normally, whereas in rab3A-deficient mice, rabphilin transport to synapses is impaired. These results show that rabphilin binds to vesicles via rab3s, consistent with an effector function of rabphilin for a synaptic rab3-signal. Surprisingly, however, no abnormalities in synaptic transmission or plasticity were observed in rabphilin-deficient mice; synaptic properties that are impaired in rab3A knock-out mice were unchanged in rabphilin knock-out mice. Our data thus demonstrate that rabphilin is endowed with the properties of a rab3 effector but is not essential for the regulatory functions of rab3 in synaptic transmission.