Bacteriophage IBB-PF7A loaded on sodium alginate-based films to prevent microbial meat spoilage

Despite the recent advances achieved in food industries to fulfil the growing consumer demand for high quality and food safety, microbial contamination remains a serious issue. This study aimed to incorporate IBB-PF7A bacteriophage (phage) onto sodium alginate-based films crosslinked with calcium chloride, to prevent poultry spoilage caused by Pseudomonas fluorescens. Films were prepared by casting and characterized in terms of phage loading, distribution, stability, release profile and antimicrobial performance. Results showed that phages were successfully incorporated as evidenced by their viability and homogeneous distribution within the films as assessed by microscopy. A decrease in phage viability was only detected after 8 weeks when stored under refrigerated conditions. Antimicrobial activity demonstrated that incorporated phages significantly impaired P. fluorescens growth. Films' antimicrobial efficacy was further demonstrated on chicken breast fillets artificially inoculated, decreasing 2Log P. fluorescens viable cell counts in the first two days and reductions were maintained up to 5 days of exposure (1 Log). These results highlight that phage incorporation onto sodium-alginate-based films constitutes a simple approach of preserving the antimicrobial activity of phages in a dried and insoluble format, that can further be applied in food industry for the prevention of microbial spoilage.AM, CM, MC acknowledge the Portuguese Foundation for Science and Technology (FCT) grants SFRH/BD/132911/2017, SFRH/BD/ 94434/2013, and SFRH/BD/122897/2016. SS is an Investigador FCT (IF/01413/2013). This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI-01-0145-FEDER-006684) and BioTecNorte operation (NORTE01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte and the Project RECI/BBB-EBI/0179/ 2012 (FCOMP-01-0124-FEDER-027462).info:eu-repo/semantics/publishedVersio

In vivo imaging of murid herpesvirus-4 infection

Luciferase-based imaging allows a global view of microbial pathogenesis. We applied this technique to gammaherpesvirus infection by inserting a luciferase expression cassette into the genome of murine herpesvirus-4 (MuHV-4). The recombinant virus strongly expressed luciferase in lytically infected cells without significant attenuation. We used it to compare different routes of virus inoculation. After intranasal infection of anaesthetized mice, luciferase was expressed in the nose and lungs for 7–10 days and in lymphoid tissue, most consistently the superficial cervical lymph nodes, for up to 30 days. Gastrointestinal infection was not observed. Intraperitoneal infection was very different to intranasal, with strong luciferase expression in the liver, kidneys, intestines, reproductive tract and spleen, but none in the nose or lungs. The nose has not previously been identified as a site of MuHV-4 infection. After intranasal infection of non-anaesthetized mice, it was the only site of non-lymphoid luciferase expression. Nevertheless, lymphoid colonization and persistence were still established, even at low inoculation doses. In contrast, virus delivered orally was very poorly infectious. Inoculation route therefore had a major impact on pathogenesis. Low dose intranasal infection without anaesthesia seems most likely to mimic natural transmission, and may therefore be particularly informative about normal viral gene functions

An In Vitro System for Studying Murid Herpesvirus-4 Latency and Reactivation

The narrow species tropisms of Epstein-Barr Virus (EBV) and the Kaposi's Sarcoma -associated Herpesvirus (KSHV) have made Murid Herpesvirus-4 (MuHV-4) an important tool for understanding how gammaherpesviruses colonize their hosts. However, while MuHV-4 pathogenesis studies can assign a quantitative importance to individual genes, the complexity of in vivo infection can make the underlying mechanisms hard to discern. Furthermore, the lack of good in vitro MuHV-4 latency/reactivation systems with which to dissect mechanisms at the cellular level has made some parallels with EBV and KSHV hard to draw. Here we achieved control of the MuHV-4 lytic/latent switch in vitro by modifying the 5′ untranslated region of its major lytic transactivator gene, ORF50. We terminated normal ORF50 transcripts by inserting a polyadenylation signal and transcribed ORF50 instead from a down-stream, doxycycline-inducible promoter. In this way we could establish fibroblast clones that maintained latent MuHV-4 episomes without detectable lytic replication. Productive virus reactivation was then induced with doxycycline. We used this system to show that the MuHV-4 K3 gene plays a significant role in protecting reactivating cells against CD8+ T cell recognition

Cross-Protection to New Drifted Influenza A(H3) Viruses and Prevalence of Protective Antibodies to Seasonal Influenza, During 2014 in Portugal

INTRODUCTION: Immune profile for influenza viruses is highly changeable over time. Serological studies can assess the prevalence of influenza, estimate the risk of infection, highlight asymptomatic infection rate and can also provide data on vaccine coverage. The aims of the study were to evaluate pre-existing cross-protection against influenza A(H3) drift viruses and to assess influenza immunity in the Portuguese population. MATERIALS AND METHODS: We developed a cross-sectional study based on a convenience sample of 626 sera collected during June 2014, covering all age groups, both gender and all administrative health regions of Portugal. Sera antibody titers for seasonal and new A(H3) drift influenza virus were evaluated by hemagglutination inhibition assay (HI). Seroprevalence to each seasonal influenza vaccine strain virus and to the new A(H3) drift circulating strain was estimated by age group, gender and region and compared with seasonal influenza-like illness (ILI) incidence rates before and after the study period. RESULTS: Our findings suggest that seroprevalences of influenza A(H3) (39.9%; 95% CI: 36.2-43.8) and A(H1)pdm09 (29.7%; 95% CI: 26.3-33.4) antibodies were higher than for influenza B, in line with high ILI incidence rates for A(H3) followed by A(H1)pdm09, during 2013/2014 season. Low pre-existing cross-protection against new A(H3) drift viruses were observed in A(H3) seropositive individuals (46%). Both against influenza A(H1)pdm09 and A(H3) seroprotection was highest in younger than 14-years old. Protective antibodies against influenza B were highest in those older than 65years old, especially for B/Yamagata lineage, 33.3% (95% CI: 25.7-41.9). Women showed a high seroprevalence to influenza, although without statistical significance, when compared to men. A significant decreasing trend in seroprotection from north to south regions of Portugal mainland was observed. CONCLUSIONS: Our results emphasize that low seroprotection increases the risk of influenza infection in the following winter season. Seroepidemiological studies can inform policy makers on the need for vaccination and additional preventive measures.info:eu-repo/semantics/publishedVersio

Murid Herpesvirus-4 Exploits Dendritic Cells to Infect B Cells

Dendritic cells (DCs) play a central role in initiating immune responses. Some persistent viruses infect DCs and can disrupt their functions in vitro. However, these viruses remain strongly immunogenic in vivo. Thus what role DC infection plays in the pathogenesis of persistent infections is unclear. Here we show that a persistent, B cell-tropic gamma-herpesvirus, Murid Herpesvirus-4 (MuHV-4), infects DCs early after host entry, before it establishes a substantial infection of B cells. DC-specific virus marking by cre-lox recombination revealed that a significant fraction of the virus latent in B cells had passed through a DC, and a virus attenuated for replication in DCs was impaired in B cell colonization. In vitro MuHV-4 dramatically altered the DC cytoskeleton, suggesting that it manipulates DC migration and shape in order to spread. MuHV-4 therefore uses DCs to colonize B cells

Global mRNA Degradation during Lytic Gammaherpesvirus Infection Contributes to Establishment of Viral Latency

During a lytic gammaherpesvirus infection, host gene expression is severely restricted by the global degradation and altered 3′ end processing of mRNA. This host shutoff phenotype is orchestrated by the viral SOX protein, yet its functional significance to the viral lifecycle has not been elucidated, in part due to the multifunctional nature of SOX. Using an unbiased mutagenesis screen of the murine gammaherpesvirus 68 (MHV68) SOX homolog, we isolated a single amino acid point mutant that is selectively defective in host shutoff activity. Incorporation of this mutation into MHV68 yielded a virus with significantly reduced capacity for mRNA turnover. Unexpectedly, the MHV68 mutant showed little defect during the acute replication phase in the mouse lung. Instead, the virus exhibited attenuation at later stages of in vivo infections suggestive of defects in both trafficking and latency establishment. Specifically, mice intranasally infected with the host shutoff mutant accumulated to lower levels at 10 days post infection in the lymph nodes, failed to develop splenomegaly, and exhibited reduced viral DNA levels and a lower frequency of latently infected splenocytes. Decreased latency establishment was also observed upon infection via the intraperitoneal route. These results highlight for the first time the importance of global mRNA degradation during a gammaherpesvirus infection and link an exclusively lytic phenomenon with downstream latency establishment

Antibody arrests γ-herpesvirus olfactory super-infection independently of neutralization

Protecting against persistent viruses is an unsolved challenge. The clearest example for a gamma-herpesvirus is resistance to super-infection by Murid herpesvirus-4 (MuHV-4). Most experimental infections have delivered MuHV-4 into the lungs. A more likely natural entry site is the olfactory epithelium. Its protection remains unexplored. Here, prior exposure to olfactory MuHV-4 gave good protection against super-infection. The protection was upstream of B cell infection, which occurs in lymph nodes, and showed redundancy between antibody and T cells. Adding antibody to virions that blocked heparan binding strongly reduced olfactory host entry - unlike in the lungs, opsonized virions did not reach IgG Fc receptor+ myeloid cells. However, the nasal antibody response to primary infection was too low to reduce host entry. Instead, the antibody acted downstream, reducing viral replication in the olfactory epithelium. This depended on IgG Fc receptor engagement rather than virion neutralization. Thus antibody can protect against natural γ-herpesvirus infection before it reaches B cells and independently of neutralization

Interpersonal coordination tendencies, decision-making and information governing dynamics in rugby union

In this chapter, ideas from ecological psychology and nonlinear dynamics are integrated to characterise decision-making as an emergent property of self-organisation processes in the interpersonal interactions that occur in sports teams. A conceptual model is proposed to capture constraints on dynamics of decisions and actions in dyadic systems, which has been empirically evaluated in simulations of interpersonal interactions in team sports. For this purpose, co-adaptive interpersonal dynamics in team sports such as rubgy union have been studied to reveal control parameter and collective variable relations in attacker-defender dyads. Although interpersonal dynamics of attackers and defenders in 1 vs 1 situations showed characteristics of chaotic attractors, the informational constraints of rugby union typically bounded dyadic systems into low dimensional attractors. Our work suggests that the dynamics of attacker-defender dyads can be characterised as an evolving sequence since players' positioning and movements are connected in diverse ways over time

Information-governing dynamics of attacker-defender interactions in youth rugby union

Previous work on dynamics of interpersonal interactions in 1 vs. 1 sub-phases of basketball has identified changes in interpersonal distance between an attacker and defender as a potential control parameter for influencing organizational states of attacker-defender dyads. Other studies have reported the constraining effect of relative velocity between an attacker and defender in 1 vs. 1 dyads. To evaluate the relationship between these candidate control parameters, we compared the impact of both interpersonal distance and relative velocity on the pattern-forming dynamics of attacker-defender dyads in the sport of rugby union. Results revealed that when interpersonal distance achieved a critical value of less than 4 m, and relative velocity values increased or were maintained above 1 m · s-1, a successful outcome (i.e. clean attempt) for an attacker was predicted. Alternatively, when values of relative velocity suddenly decreased below this threshold, at the same critical value of interpersonal distance, a successful outcome for the defender was predicted. Data demonstrated how the coupling of these two potential, nested control parameters moved the dyadic system to phase transitions, characterized as a try or a tackle. Observations suggested that relative velocity increased its influence on the organization of attacker-defender dyads in rugby union over time as spatial proximity to the try line increased