EMS Access Constraints And Response Time Delays For Deprived Critically Ill Patients Near Paris, France.

Increased emergency medical services (EMS) response times and areas of low socioeconomic status are both associated with poorer outcomes for several time-sensitive medical conditions attended to by medical personnel before a patient is hospitalized. We evaluated the association between EMS response times, area deprivation level, and on-scene access constraints encountered by EMS in a large urban area in France. We conducted a multicenter prospective cohort study of EMS dispatches occurring in the forty-seven cities in a region southeast of Paris. We fit multilevel mixed-effects linear regression models for multivariate assessment of the predictors of EMS response times and then used multivariate logistic regression on outcomes among a subgroup of patients presenting with out-of-hospital cardiac arrest. We found evidence that access constraints were more frequently encountered by EMS in the most deprived areas compared to less deprived ones, and were associated with increased EMS response times until patient contact and with poorer outcomes from cardiac arrest. Strategies to anticipate and overcome access constraints should be implemented to improve outcomes for emergent conditions attended to by prehospital medical teams.

Oxygen and viruses: a breathing story.

The effect of oxygen on virus replication is complex, and the role of hypoxia-inducible factor 1α (HIF-1α) in the metabolism of virus-infected cells remains uncertain. Solid tumours are hypoxic, and some viruses use this low oxygen tension level to facilitate their replication in tumour cells, thereby causing cell lysis. In addition, the interactions between viruses and HIF-1α may stimulate a trained immunity. However, the evolutionary basis for the oxygen regulatory mechanism of virus replication is ill-defined and requires further investigation.

Expression of VP7, a Bluetongue virus group specific antigen by viral vectors: analysis of the induced immune responses and evaluation of protective potential in sheep.

Bluetongue virus (BTV) is an economically important Orbivirus transmitted by biting midges to domestic and wild ruminants. The need for new vaccines has been highlighted by the occurrence of repeated outbreaks caused by different BTV serotypes since 1998. The major group-reactive antigen of BTV, VP7, is conserved in the 26 serotypes described so far, and its role in the induction of protective immunity has been proposed. Viral-based vectors as antigen delivery systems display considerable promise as veterinary vaccine candidates. In this paper we have evaluated the capacity of the BTV-2 serotype VP7 core protein expressed by either a non-replicative canine adenovirus type 2 (Cav-VP7 R0) or a leporipoxvirus (SG33-VP7), to induce immune responses in sheep. Humoral responses were elicited against VP7 in almost all animals that received the recombinant vectors. Both Cav-VP7 R0 and SG33-VP7 stimulated an antigen-specific CD4+ response and Cav-VP7 R0 stimulated substantial proliferation of antigen-specific CD8+ lymphocytes. Encouraged by the results obtained with the Cav-VP7 R0 vaccine vector, immunized animals were challenged with either the homologous BTV-2 or the heterologous BTV-8 serotype and viral burden in plasma was followed by real-time RT-PCR. The immune responses triggered by Cav-VP7 R0 were insufficient to afford protective immunity against BTV infection, despite partial protection obtained against homologous challenge. This work underscores the need to further characterize the role of BTV proteins in cross-protective immunity.

Functional balance between the hemagglutinin and neuraminidase of influenza A(H1N1)pdm09 HA D222 variants.

D222G/N substitutions in A(H1N1)pdm09 hemagglutinin may be associated with increased binding of viruses causing low respiratory tract infections and human pathogenesis. We assessed the impact of such substitutions on the balance between hemagglutinin binding and neuraminidase cleavage, viral growth and in vivo virulence.Seven viruses with differing polymorphisms at codon 222 (2 with D, 3 G, 1 N and 1 E) were isolated from patients and characterized with regards hemagglutinin binding affinity (Kd) to α-2,6 sialic acid (SAα-2,6) and SAα-2,3 and neuraminidase enzymatic properties (Km, Ki and Vmax). The hemagglutination assay was used to quantitatively assess the balance between hemagglutinin binding and neuraminidase cleavage. Viral growth properties were compared in vitro in MDCK-SIAT1 cells and in vivo in BALB/c mice. Compared with D222 variants, the binding affinity of G222 variants was greater for SAα-2,3 and lower for SAα-2,6, whereas that of both E222 and N222 variants was greater for both SAα-2,3 and SAα-2,6. Mean neuraminidase activity of D222 variants (16.0 nmol/h/10(6)) was higher than that of G222 (1.7 nmol/h/10(6) viruses) and E/N222 variants (4.4 nmol/h/10(6) viruses). The hemagglutination assay demonstrated a deviation from functional balance by E222 and N222 variants that displayed strong hemagglutinin binding but weak neuraminidase activity. This deviation impaired viral growth in MDCK-SIAT1 cells but not infectivity in mice. All strains but one exhibited low infectious dose in mice (MID50) and replicated to high titers in the lung; this D222 strain exhibited a ten-fold higher MID50 and replicated to low titers. Hemagglutinin-neuraminidase balance status had a greater impact on viral replication than hemagglutinin affinity strength, at least in vitro, thus emphasizing the importance of an optimal balance for influenza virus fitness. The mouse model is effective in assessing binding to SAα-2,3 but cannot differentiate SAα-2,3- from SAα-2,6- preference, nor estimate the hemagglutinin-neuraminidase balance in A(H1N1)pdm09 strains.

Production and purification of non replicative canine adenovirus type 2 derived vectors.

Adenovirus (Ad) derived vectors have been widely used for short or long-term gene transfer, both for gene therapy and vaccine applications. Because of the frequent pre-existing immunity against the classically used human adenovirus type 5, canine adenovirus type 2 (CAV2) has been proposed as an alternative vector for human gene transfer. The well-characterized biology of CAV2, together with its ease of genetic manipulation, offer major advantages, notably for gene transfer into the central nervous system, or for inducing a wide range of protective immune responses, from humoral to cellular immunity. Nowadays, CAV2 represents one of the most appealing nonhuman adenovirus for use as a vaccine vector. This protocol describes a simple method to construct, produce and titer recombinant CAV2 vectors. After cloning the expression cassette of the gene of interest into a shuttle plasmid, the recombinant genomic plasmid is obtained by homologous recombination in the E. coli BJ5183 bacterial strain. The resulting genomic plasmid is then transfected into canine kidney cells expressing the complementing CAV2-E1 genes (DK-E1). A viral amplification enables the production of a large viral stock, which is purified by ultracentrifugation through cesium chloride gradients and desalted by dialysis. The resulting viral suspension routinely has a titer of over 10(10) infectious particles per ml and can be directly administrated in vivo.

Genetic content of Influenza H3N2 vaccine seeds.

Influenza vaccine seeds produced in chicken eggs are selected through HA and NA surface glycoproteins antigenicity, as well as through high replicative ability. Here we characterize the genetic content of recently used thirteen H3N2 influenza vaccine seeds. Interestingly, sequence analysis of the vaccine seeds shows reassortment events leading to PR8:H3N2 segment constellations, ranging from the 6:2 to 2:6 constellations. This study shows that the H3N2 PB1 is the most frequent internal segment incorporated in the tested vaccines seeds.