Motorcycling performance and sleepiness during an extended ride on a dynamic simulator: relationship with stress biomarkers.

OBJECTIVE: Powered two-wheelers (PTW) make up a large proportion of fatal accidents. The aim of this study was to investigate the effects of time-of-day and total sleep deprivation (SD) on simulated motorcycling performance during extended riding sessions (60 min), while evaluating stress mechanisms. APPROACH: A total of 16 healthy males participated in four simulated motorcycling sessions at 07:00, 11:00, 15:00 and 19:00, including city (8 min), country (2 min) and highway pathways (40 min), after a normal night of sleep and after total SD (30 h), in a randomized counterbalanced order. The recorded motorcycle parameters included: variation of lateral position, number of inappropriate line crossings (ILC), falls, riding errors, speed and speed limit violations. Subject parameters included the number of microsleeps in each pathway, the number of lapses during the 3-min psychomotor vigilance task (PVT-Brief version), and the Karolinska sleepiness scale (KSS) score. Saliva samples were used to assess cortisol (sC), α-amylase (sAA), and chromogranin-A (sCgA). ANOVAs and Pearson's correlation analysis were performed between these variables. MAIN RESULTS: Most parameters were influenced by an interaction effect between 'Motorcycling pathways' × 'SD' (speed (p < 0.05), legal speed violations (p < 0.01), variation of lateral position (p < 0.001), falls (p < 0.001), EEG-microsleeps (p < 005)). An interaction effect between 'SD' × 'Time-of-day' influenced the number of ILCs (p < 0.01), sC (p < 0.05) and sCgA (p < 0.05) levels. SD affected KSS scores (p < 0.001) and PVT lapses (p < 0.05). The highest disturbances were associated with highway motorcycling simulation. SIGNIFICANCE: Sleepiness due to circadian or SD and fatigue effects significantly affect riding and increase the risks involved with PTWs. The activation of both stress systems seems not sufficient to alleviate these deleterious effects.

Pulsed-plasma gas-discharge inactivation of microbial pathogens in chilled poultry wash water.

A pulsed-plasma gas-discharge (PPGD) system was developed for the novel decontamination of chilled poultry wash water. Treatment of poultry wash water in the plasma generation chamber for up to 24 s at 4 degrees C reduced Escherichia coli NCTC 9001, Campylobacter jejuni ATCC 33560, Campylobacter coli ATCC 33559, Listeria monocytogenes NCTC 9863, Salmonella enterica serovar Enteritidis ATCC 4931, and S. enterica serovar Typhimurium ATCC 14028 populations to non-detectable levels (< or = 8 log CFU/ml). Although similar PPGD treatments at 4 degrees C also produced significant reductions (> or = 3 log CFU/ml) in recalcitrant B. cereus NCTC 11145 endospore numbers within 30 s, the level of endospore reduction was dependent on the nature of the sparged gas used in the plasma treatments. Scanning electron microscopy revealed that significant damage occurred at the cellular level in PPGD-treated test organisms. This electrotechnology delivers energy in intense ultrashort bursts, generating products such as ozone, UV light, acoustic and shock waves, and pulsed electric fields that have multiple bactericidal properties. This technology offers an exciting complementary or alternative approach for treating raw poultry wash water and for preventing cross-contamination in processing environments.

Reliability of simulator driving tool for evaluation of sleepiness, fatigue and driving performance.

STUDY OBJECTIVE: To compare the impact of extended wakefulness (i.e., sleepiness) and prolonged driving (i.e., fatigue) at the wheel in simulated versus real-life driving conditions. DESIGN: Participants drove on an INRETS-MSIS SIM2 simulator in a research laboratory or an open French highway during 3 nocturnal driving sessions. A dose-response design of duration of nocturnal driving was used: a 2 h short driving session (3-5 AM), a 4 h intermediate driving session (1-5 AM) and an 8 h long driving session (9 PM-5 AM). PARTICIPANTS: Two groups of healthy male drivers (20 for simulated driving and 14 drivers for real driving; mean age±SD=22.3±1.6 years), free of sleep disorders. MEASUREMENTS: Number of inappropriate line crossings, self-rated fatigue and sleepiness were recorded in the last hour of driving sessions to control the effects of prior waking time and time of day. RESULTS: Compared to the daytime reference session, both simulated and real driving performance were affected by a short nocturnal driving session (P<.05 and P<.001, respectively). Extension of nocturnal driving duration affected simulated performance nonlinearly and more severely than that of real driving (P<.001). Compared to the daytime reference session, short nocturnal simulated and real driving sessions increased self-perceived fatigue and sleepiness. Real and simulated driving conditions had an identical impact on fatigue and sleepiness during extended periods of nocturnal driving. CONCLUSIONS: In healthy subjects, the INRETS-MSIS SIM2 simulator appropriately measures driving impairment in terms of inappropriate line crossings related to extended wakefulness but has limitations to measure the impact of extended driving on drivers' performance.

Evidence of lethal and sublethal injury in food-borne bacterial pathogens exposed to high-intensity pulsed-plasma gas discharges.

AIMS: To apply scanning electron microscopy, image analysis and a fluorescent viability stain to assess lethal and sublethal injury in food-borne bacteria exposed to pulsed-plasma gas discharges (PPGD). METHODS AND RESULTS: The fluorescent redox probe 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) was used for enumerating actively respiring cells of Campylobacter jejuni, Escherichia coli, Listeria monocytogenes, Staphylococcus aureus and Salmonella enterica serovar Typhimurium that were suspended in sterile water at 4 degrees C and exposed to separate PPGD and heat treatments. While there was good agreement between use of respiratory staining (RS) and direct-selective agar plate counting (PC) for enumerating untreated bacteria, there were c. 1 and 3 log-unit differences in surviving cell numbers per millilitre for test organisms subjected to PPGD and heat treatments respectively, when enumerated by these different viability indicators. PPGD-treated bacteria were markedly altered at the cellular level when examined by scanning electron microscopy. CONCLUSIONS: Use of this RS method revealed that substantial subpopulations of test bacteria rendered incapable of forming colonies by separate PPGD and heat treatments may remain metabolically active. SIGNIFICANCE AND IMPACT OF THE STUDY: Use of this RS method offers interesting perspectives on assessing established and novel microbial inactivation methods, and may also provide a better understanding of mechanisms involved in microbial inactivation induced by high-intensity PPGD treatments.

Human anti-mouse xenorecognition. Provision of noncognate interactions reveals plasticity of T cell repertoire.

DAP.3 transfectants expressing native H-2E molecules with or without human LFA-3 and ICAM-1 (intercellular adhesion molecule-1) failed to induce proliferation by human peripheral blood T cells. Introduction of sequence from the DR beta 2 domain into the H-2E molecule led to the induction of detectable proliferation, which was substantially augmented by co-expression of human LFA-3 and ICAM-1 to levels comparable to those induced by DAP.3 cells co-expressing wild-type DR alloantigens with human LFA-3/ICAM-1. In marked contrast, cells expressing native H-2A molecules together with human accessory molecules provoked strong primary proliferative responses. The results of Ab inhibition experiments confirmed that this was caused by direct xenorecognition. In limiting dilution assays the frequency of anti-H-2A, IL-2-secreting, CD4+ human T cells was only fivefold lower than that measured against a DR alloantigen expressed on the same background. No measurable frequency was recorded against H-2E-expressing cells. Evidence to suggest that this difference was a result of isotype-specific differences in the interaction with CD4 was provided using transfectants expressing DR alloantigens with either the H-2E or H-2A beta 2 domain. DR molecules with the H-2A beta 2 domain stimulated a substantially stronger response than those with the H-2E beta 2 domain. These results challenge the view that xenogeneic T cell responses between evolutionarily distant species are weak; further emphasize the influence of the interaction between the T cell co-receptor molecule CD4, with its MHC class II molecular ligand on the strength of primary xenoresponses; and suggest that MHC class II isotypes may differ substantially in their interaction with CD4.

Conserved transmembrane tyrosine residues of the TCR beta chain are required for TCR expression and function in primary T cells and hybridomas.

The T cell receptor (TCR) beta chain transmembrane domain contains two evolutionarily conserved tyrosines (Y). In this study, the functional basis for the evolutionary conservation is addressed by mutation of the residues, expression of the mutants in hybridoma and primary T cells, and examination of TCR signaling function. We find that the phenotype of the mutants, both surface expression and ability to signal for IL-2 production, is highly variable in different mouse T hybridoma lines. Although we have not been able to determine the basis for these differences in the hybridomas, expression of the mutants in primary T cells provides a definitive assessment of mutant phenotype. We show that mutation of the N-terminal Y to either leucine (L) or alanine (A) results in low surface expression in primary T cells, while mutation of both N- and C-terminal Y to A or L abrogates surface expression. However, the more conservative mutation of both transmembrane Y to phenylalanine maintained receptor surface expression and assembly while severely disrupting signaling in primary T cells. Our data demonstrate that TCR beta chain transmembrane Y are essential for TCR signal transduction as well as complex assembly. These findings suggest that protein-protein interactions involving membrane-spanning domains are likely relevant for TCR signal transduction mechanisms.

Quantitative but not qualitative variation in MHC class II alters CD4 interaction and influences T cell repertoire formation.

The influence of the interaction between CD4 and MHC class II molecules on selection of the T cell repertoire was studied in transgenic mice expressing human or human/mouse hybrid MHC class II beta chains. Either wild-type DR beta chains (DR1 beta) or hybrid beta chains comprising the beta1 domain of DR and the beta2, transmembrane, and intracytoplasmic domains of I-E (DRbeta 1Ebeta2) were introduced into and expressed in transgenic mice as a heterodimer with endogenous I-E alpha. Mice expressing low levels of DR1beta:I-E alpha or those expressing low or higher levels of the hybrid DRbeta 1Ebeta2:I-E alpha were studied. Immunization with a suboptimal dose of influenza nucleoprotein peptide exposed a fivefold lower frequency of DR-restricted, peptide-specific, IL-2-secreting T cells in the mice with low-level expression of DRbeta1 Ebeta2:I-E alpha when compared to mice expressing the same molecule at higher levels. The frequency in DRbeta wild-type mice was only twofold lower than that measured in mice with comparable levels of expression of DRbeta 1Ebeta2. These results suggest that positive selection is sensitive to quantitative variation in MHC class II density, unmasked when antigen is limiting, but is relatively insensitive to qualitative variation in the MHC class II: CD4 interaction.

Transmembrane polar residues of TCR beta chain are required for signal transduction.

Mutagenic analyses have identified structural motifs important for TCR-mediated signaling in the antigen-binding chains, CD3 and zeta subunits of the TCR complex. In this study, we altered selected residues in the transmembrane and extracellular constant regions of the TCR beta chain and expressed the mutants in a T hybridoma line bearing endogenous receptor. We measured cytokine production and apoptosis in response to antigen or antibody. We found that mutation of one or both of the transmembrane tyrosine residues in the TCR beta chain caused a marked reduction in responsiveness. Mutation of the transmembrane serine to alanine also reduced responses, although less markedly. Immunoprecipitation analyses showed that the TCR beta mutations did not alter association with zeta. These experiments identify a signaling role for the transmembrane domain of the TCR beta chain.

Antigen presentation by T cells inhibits IL-2 production and induces IL-4 release due to altered cognate signals.

Conflicting results of the effects of Ag presentation by MHC class II-expressing T cells have been described. In some studies class II-expressing T cells have been shown to act as effective APCs, while others have reported that the recognition of Ag on the surface of another T cell inactivates IL-2 production. In this study we have investigated the mechanisms involved in Ag presentation by T cells. The results obtained suggest that 1) lack of costimulation is not responsible for the inhibitory effects of T cell Ag presentation on IL-2 production; the provision of costimulation by immobilized anti-CD28 Ab or by the addition of accessory cells failed to reverse the effects of T cell Ag presentation, but restored the response to immobilized anti-CD3; 2) T cell Ag presentation induced a minimal increase in intracellular Ca2+ compared with that induced by antigen-pulsed B cells; this difference in the calcium response is not explained by quantitative differences in ligand density between B cells and T cells; and 3) despite the weak calcium signal, T cell presentation supported IL-4 release in the absence of IL-2 production. Taken together these data suggest that T cell Ag presentation leads to altered TCR/CD3-transduced signals, which biases the T cell towards a Th2 phenotype.