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Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a novel coronavirus that has given rise to the devastating global pandemic. In most cases, SARS-CoV-2 infection results in the development of viral pneumonia and acute respiratory distress syndrome, known as “coronavirus disease 2019” or COVID-19. Intriguingly, besides the respiratory tract, COVID-19 affects other organs and systems of the human body. COVID-19 patients with pre-existing cardiovascular disease have a higher risk of death, and SARS-CoV-2 infection itself may cause myocardial inflammation and injury. One possible explanation of such phenomena is the fact that SARS-CoV-2 utilizes angiotensin-converting enzyme 2 (ACE2) as the receptor required for viral entry. ACE2 is expressed in the cells of many organs, including the heart. ACE2 functions as a carboxypeptidase that can cleave several endogenous substrates, including angiotensin II, thus regulating blood pressure and vascular tone. It remains largely unknown if the SARS-CoV-2 infection alters the enzymatic properties of ACE2, thereby contributing to cardiovascular complications in patients with COVID-19. Here, we demonstrate that ACE2 cleavage of des-Arg9-bradykinin substrate analog is markedly accelerated, while cleavage of angiotensin II analog is minimally affected by the binding of spike protein. These findings may have implications for a better understanding of COVID-19 pathogenesis., Graphical Abstract Graphical Abstract more...

Sindbis virus causes age-dependent encephalomyelitis in mice. Young mice and immature neurons replicate the virus to high titers and die from infection while older mice and mature neurons restrict replication and survive infection. Studies to identify factors that affect maturation-dependent virus replication in neurons have used AP-7 rat olfactory neuronal cells that can be differentiated in vitro, and have identified roles for host transcription factors interferon regulatory protein (IRF) 7 and NF-B and viral proteins E2 and nsP3. IRF7 is required for neuronal survival and deficiency leads to paralysis and death of 4–6 weeks old C57BL/6 mice. Activation of NF-κB works in conjunction with PKR to facilitate replication in mature neurons by promoting shut-off of host protein synthesis and increasing translation of the viral structural proteins from subgenomic RNA. The macro domain of nsP3 binds and hydrolyses ADP-ribose from ADP-ribosylated proteins and is important for initiation of neuronal infection and for synthesis of viral structural proteins. Thus, neurons regulate translation of the structural proteins from subgenomic RNA required for the production of the infectious virus. more...

Interferon (IFN) regulatory factors (IRFs) are important determinants of the innate response to infection. We evaluated the role(s) of combined and individual IRF deficiencies in the outcome of infection of C57BL/6 mice with Sindbis virus, an alphavirus that infects neurons and causes encephalomyelitis. The brain and spinal cord levels of Irf7, but not Irf3 mRNAs, were increased after infection. IRF3/5/7-/- and IRF3/7-/- mice died within 3-4 days with uncontrolled virus replication, similar to IFNα receptor-deficient mice, while all wild-type (WT) mice recovered. IRF3-/- and IRF7-/- mice had brain levels of IFNα that were lower, but brain and spinal cord levels of IFNβ and IFN-stimulated gene mRNAs that were similar to or higher than WT mice without detectable serum IFN or increases in Ifna or Ifnb mRNAs in the lymph nodes, indicating that the differences in outcome were not due to deficiencies in the central nervous system (CNS) type I IFN response. IRF3-/- mice developed persistent neurological deficits and had more spinal cord inflammation and higher CNS levels of Il1b and Ifnγ mRNAs than WT mice, but all mice survived. IRF7-/- mice died 5-8 days after infection with rapidly progressive paralysis and differed from both WT and IRF3-/- mice in the induction of higher CNS levels of IFNβ, tumour necrosis factor (TNF) α and Cxcl13 mRNA, delayed virus clearance and more extensive cell death. Therefore, fatal disease in IRF7-/- mice is likely due to immune-mediated neurotoxicity associated with failure to regulate the production of inflammatory cytokines such as TNFα in the CNS. more...

The melting behavior of nascent poly(tetrafluoroethylene) (PTFE) was investigated by way of differential scanning calorimetry (DSC). It is well known that the melting temperature of nascent PTFE is about 344∘ C, but reduces to 327∘ C for once molten material. In this study, the melting temperature of nascent PTFE crystals was found to strongly depend on heating rate, decreasing considerably for slow heating rates. In addition, during isothermal experiments in the temperature range of 327∘C, Polymers, 12 (4), ISSN:2073-4360 more...

While crystallization behavior of isotactic polypropylene homopolymers had been subject to a wide range of experimental and modeling studies, this is not the case for propylene-ethylene random copolymers (PPR). This class of polymers offers up to now significant challenges, both from an experimental as well as a modeling perspective. The ethylene incorporation in the propylene chains, as well as the distribution of this comonomer, has a marked effect on the crystallization kinetics. Moreover, the presence of these defects causes a clear separation between primary crystallization (i.e. space filling) and subsequent secondary crystallization (increase of crystallinity in filled space) within the spherulitic skeletons, particularly subsequent at high primary crystallization temperatures. In this work, the underlying mechanism is first quantified by means of a combination of in-situ WAXD and SAXS experiments, as well as ex-situ WAXD experiments and calorimetric measurements. Based on these experiments an extended model framework is presented, capable of predicting multiphase non-isothermal crystallization kinetics as well as the final crystallinity as a function of the applied thermal conditions relevant for processing. The chemical composition distribution (CCD) of the ethylene comonomer serves as critical input to parameterize the model. Optical microscopy- and DSC experiments are used for parameterization of the primary crystallization model. The model developed in this study is, in principle, applicable to all polypropylenes, ranging from homo-polymers to random copolymers with variable comonomer content and/or CCD but, so far, only applied and validated on one PPR. To validate the model and the parameters for a given PPR, several non-isothermal and isothermal experiments (the latter followed by subsequent cooling) are conducted over a wide range of crystallization temperatures and cooling rates. The good match between experiments and model predictions demonstrates the power of the newly developed framework. The final crystallinity, the amount of α- and γ-phase, and the ratio between primary and secondary crystallization can be predicted as a function of the time-temperature history. To the best knowledge of the authors, it is the first time that such a direct connection with the CCD is incorporated in a crystallization model. Consequently, the model offers a new tool to bridge the gap between chemical structure and resulting product properties, which now has come one step closer for PPR systems. more...

Central nervous system (CNS) infection of C57BL/6 mice with the TE strain of Sindbis virus (SINV) provides a valuable animal model for studying the pathogenesis of alphavirus encephalomyelitis. While SINV TE inoculated intracranially causes little mortality, 20–30 % of mice inoculated intranasally (IN) died 8 to 11 days after infection, the period during which immune cells typically infiltrate the brain and clear infectious virus. To examine the mechanism behind the mortality, mice infected IN with SINV TE were monitored for evidence of neurological disease, and those with signs of severe disease (moribund) were sacrificed and tissues collected. Mice showing the usual mild signs of encephalomyelitis were concurrently sacrificed to serve as time-matched controls (sick). Sixty-eight per cent of the moribund mice, but none of the sick mice, showed upper gastrointestinal bleeding due to gastric ulceration. Clinical disease and gastrointestinal pathology could not be attributed to direct viral infection of tissues outside of the CNS, and brain pathology and inflammation were comparable in sick and moribund mice. However, more SINV antigen was present in the brains of moribund mice, and clearance of infectious virus from the CNS was delayed compared to sick mice. Lower levels of SINV-specific IgM and fewer B220+ B cells were present in the brains of moribund mice compared to sick mice, despite similar levels of antiviral IgM and IgG in serum. These findings highlight the importance of the local antibody response in determining the outcome of viral encephalomyelitis and offer a model system for understanding individual variation in this response. more...

The performance of polypropylene-poly(ethylene brassylate) block and graft copolymers and a polypropylene-polycaprolactone graft copolymer as compatibilizers for polypropylene-rich polypropylene/bisphenol A polycarbonate (PP/PC, 80/20 wt/wt) blends was elucidated. The copolymers were synthesized either by metal-catalyzed ring-opening polymerization or transesterification of a presynthesized polyester, initiated by hydroxyl-functionalized PPs, which themselves were obtained by catalytic routes or reactive extrusion, respectively. Spectroscopic fingerprints of the copolymers from liquid-state nuclear magnetic resonance (NMR) in combination with scanning electron microscopy (SEM), transmission electron microscopy (TEM), atomic force microscopy (AFM), dynamic mechanical thermal analysis (DMTA), and rheology analyses of the blends indicated that the compatibilizers spontaneously organize at the interface of the two immiscible polymers leading to the formation of uniform, stable, nanophase morphologies. The effect of the compatibilizers on the performance of the PP/PC blends was evaluated, and well-compatibilized PP/PC blends showed improved melt strength and strain hardening when compared to pure PP. This was verified by the successful foam extrusion using isobutane as a blowing agent of well-compatibilized PP/PC blends to low-density PP-based foams, for which normally long-chain branched PP is required. more...

Yeast prions are protein-based genetic elements that propagate through cell populations via cytosolic transfer from mother to daughter cell. Molecular chaperone proteins including Hsp70, the Hsp40/J-protein Sis1, and Hsp104 are required for continued prion propagation, however the specific requirements of chaperone proteins differ for various prions. We recently reported that Swa2, the yeast homolog of the mammalian protein auxilin, is specifically required for the propagation of the prion [URE3].1 [URE3] propagation requires both a functional J-domain and the tetratricopeptide repeat (TPR) domain of Swa2, but does not require Swa2 clathrin binding. We concluded that the TPR domain determines the specificity of the genetic interaction between Swa2 and [URE3], and that this domain likely interacts with one or more proteins with a C-terminal EEVD motif. Here we extend that analysis to incorporate additional data that supports this hypothesis. We also present new data eliminating Hsp104 as the relevant Swa2 binding partner and discuss our findings in the context of other recent work involving Hsp90. Based on these findings, we propose a new model for Swa2's involvement in [URE3] propagation in which Swa2 and Hsp90 mediate the formation of a multi-protein complex that increases the number of sites available for Hsp104 disaggregation. more...

Polychlorinated biphenyls (PCBs) are recognised reproductive and immune system toxicants in marine mammals mediated by endocrine-disrupting mechanisms. As with other predators, seals are exposed to elevated bioaccumulated concentrations of PCBs and other persistent organic pollutants (POPs). Cryopreserved plasma samples from adult ringed (Phoca hispida; n = 39) and grey (Halichoerus grypus; n = 38) seals, sampled between 1998 and 2002 from Baltic Sea, Svalbard, and Sable Island (Canada) were used to investigate relationships between PCB exposure and sex hormone concentrations (progesterone; P4, 17α-hydroxy progesterone; 17α-OH-P4, testosterone; T4, 17β-estradiol; E2, estrone; E3). Immunoassay methods were used for quantification of analytes due to the limited sample volumes available. PCB concentrations were found to be significantly higher in Baltic seals than other sampling locations and were classed as “Exposed” seals while Svalbard and Sable Is seal were classed “Reference” seals (sexes and species separate). Mean hormone concentrations in Exposed seal were lower than Reference seals, and this was statistically significantly for 17α-OH-P4 (both sexes and both species), E2 (ringed and grey seal females), and E3 (grey seal females). Regression analyses (PCB v hormone concentrations) for each sex and species revealed significant correlations for P4 (Sable Is. female grey seals and female ringed seals), 17α-OH-P4 (Sable Is. male grey seals and Svalbard male ringed seals), T4 (Svalbard male ringed seals), E2 (female ringed seals), and E3 (female ringed seals and Baltic female grey seals). Although significant correlations are not evidence of cause and effect, the potential impact of hormone changes on endocrine homeostasis and reproductive health for seal populations warrants further investigation given that PCB concentrations found here are in the same range as those currently reported in seals from these populations. more...

Polymer, 222, ISSN:0032-3861, ISSN:1873-2291

Innovation of the Ziegler–Natta polymerization process for polymerization of propene that allows tailoring of the mechanical properties of isotactic polypropylene (iPP) directly in the polymerization stage is presented. The catalyst is modified by prepolymerization of trimethylallylsilane or vinylcyclohexane so that the catalyst particles are coated by a thin skin of poly(trimethylallylsilane) or poly(vinylcyclohexane) that will act as a nucleating agent for the crystallization of iPP. The presence of the nucleating agent accelerates the crystallization of iPP and affords crystallization of the α form even upon fast crystallization by quenching the melt, a condition that generally produces crystallization of the mesomorphic form. Crystals of the α form so obtained show a nodular morphology and the absence of the spherulitic superstructure. This novel iPP material is characterized by outstanding and unexpected properties of high mechanical strength and modulus and contemporarily high ductility, flexibility and good transparency due to the nodular morphology of the α form. more...

In the summer of 2007, a wilt of endive (Cichorium endivia) cv. Myrna plants was observed on several commercial farms near Alessandria in northern Italy. Approximately 40% of the plants on each farm were symptomatic. Affected plants were stunted and yellow and their root systems were poorly developed. Basal leaves eventually wilted. Black streaks were observed in the proximal part of the leaf vascular system. Fusarium oxysporum was consistently isolated from symptomatic vascular tissue on a Fusarium-selective medium (1). To confirm identification of the pathogen, the translation elongation factor 1α and the ribosomal DNA intergenic spacer (IGS) region were partially sequenced (2). Genomic DNA was extracted from mycelia growing on potato dextrose agar. Amplification of the EF-1α region generated a sequence of 648 bp; the IGS amplicon was 2,500 bp. The EF-1α sequence (GenBank Accession No GQ398152) was 99% similar to the sequence of a F. oxysporum strain isolated from soil and a strain pathogenic on cotton plants (GenBank Accession No. EU246574). The IGS sequence (GenBank Accession No GQ398153) was 97% similar to the sequence of a F. oxysporum strain (GenBank Accession No. EF661647). Pathogenicity tests were conducted on 15-day-old endive plants (cv. Myrna). Ten plants were inoculated by dipping their roots in a 1 × 106 CFU/ml suspension of one of the isolates recovered from a wilted endive plant. Inoculated and noninoculated plants were transplanted into pots filled with steamed soil and maintained in a glasshouse at 23 to 28°C. Wilt symptoms and vascular discoloration of the roots, crown, and veins developed 60 days after inoculation. F. oxysporum was consistently reisolated from infected plants. Noninoculated plants remained healthy. The pathogenicity test was conducted twice. To our knowledge, this is the first report of wilt caused by F. oxysporum on endive in Italy. References: (1) H. Komada. Rev. Plant Prot. Res. 8:114, 1975. (2) G. Mbofung et al. Phytopathology 97:87, 2007. more...

Central nervous system (CNS) infection of C57BL/6 mice with the TE strain of Sindbis virus (SINV) provides a valuable animal model for studying the pathogenesis of alphavirus encephalomyelitis. While SINV TE inoculated intracranially causes little mortality, 20–30 % of mice inoculated intranasally (IN) died 8 to 11 days after infection, the period during which immune cells typically infiltrate the brain and clear infectious virus. To examine the mechanism behind the mortality, mice infected IN with SINV TE were monitored for evidence of neurological disease, and those with signs of severe disease (moribund) were sacrificed and tissues collected. Mice showing the usual mild signs of encephalomyelitis were concurrently sacrificed to serve as time-matched controls (sick). Sixty-eight per cent of the moribund mice, but none of the sick mice, showed upper gastrointestinal bleeding due to gastric ulceration. Clinical disease and gastrointestinal pathology could not be attributed to direct viral infection of tissues outside of the CNS, and brain pathology and inflammation were comparable in sick and moribund mice. However, more SINV antigen was present in the brains of moribund mice, and clearance of infectious virus from the CNS was delayed compared to sick mice. Lower levels of SINV-specific IgM and fewer B220(+) B cells were present in the brains of moribund mice compared to sick mice, despite similar levels of antiviral IgM and IgG in serum. These findings highlight the importance of the local antibody response in determining the outcome of viral encephalomyelitis and offer a model system for understanding individual variation in this response. more...

Sindbis virus (SINV) infection of neurons in the brain and spinal cord in mice provides a model system for investigating recovery from encephalomyelitis and antibody-mediated clearance of virus from the central nervous system (CNS). To determine the roles of IgM and IgG in recovery, we compared the responses of immunoglobulin-deficient activation-induced adenosine deaminase-deficient (AID −/− ), secretory IgM-deficient (sIgM −/− ), and AID −/− sIgM −/− double-knockout (DKO) mice with those of wild-type (WT) C57BL/6 mice for disease, clearance of infectious virus and viral RNA from brain and spinal cord, antibody responses, and B cell infiltration into the CNS. Because AID is essential for immunoglobulin class switch recombination and somatic hypermutation, AID −/− mice produce only germ line IgM, while sIgM −/− mice secrete IgG but no IgM and DKO mice produce no secreted immunoglobulin. After intracerebral infection with the TE strain of SINV, most mice recovered. Development of neurologic disease occurred slightly later in sIgM −/− mice, but disease severity, weight loss, and survival were similar between the groups. AID −/− mice produced high levels of SINV-specific IgM, while sIgM −/− mice produced no IgM and high levels of IgG2a compared to WT mice. All mice cleared infectious virus from the spinal cord, but DKO mice failed to clear infectious virus from brain and had higher levels of viral RNA in the CNS late after infection. The numbers of infected cells and the amount of cell death in brain were comparable. We conclude that antibody is required and that either germ line IgM or IgG is sufficient for clearance of virus from the CNS. IMPORTANCE Mosquito-borne alphaviruses that infect neurons can cause fatal encephalomyelitis. Recovery requires a mechanism for the immune system to clear virus from infected neurons without harming the infected cells. Antiviral antibody has previously been shown to be a noncytolytic means for alphavirus clearance. Antibody-secreting cells enter the nervous system after infection and produce antiviral IgM before IgG. Clinical studies of human viral encephalomyelitis suggest that prompt production of IgM is associated with recovery, but it was not known whether IgM is effective for clearance. Our studies used mice deficient in production of IgM, IgG, or both to characterize the antibody necessary for alphavirus clearance. All mice developed similar signs of neurologic disease and recovered from infection. Antibody was necessary for virus clearance from the brain, and either early germ line IgM or IgG was sufficient. These studies support the clinical observation that prompt production of antiviral antibody is a determinant of outcome. more...

Summary Yeast prions require a core set of chaperone proteins including Sis1, Hsp70 and Hsp104 to generate new amyloid templates for stable propagation, yet emerging studies indicate that propagation of some prions requires additional chaperone activities, demonstrating chaperone specificity beyond the common amyloid requirements. To comprehensively assess such prion-specific requirements for the propagation of the [URE3] prion variant [URE3-1], we screened 12 yeast cytosolic J-proteins, and here we report a novel role for the J-protein Swa2/Aux1. Swa2 is the sole yeast homolog of the mammalian protein auxilin, which, like Swa2, functions in vesicle-mediated endocytosis by disassembling the structural lattice formed by the protein clathrin. We found that, in addition to Sis1, [URE3-1] is specifically dependent upon Swa2, but not on any of the 11 other J-proteins. Further, we show that [URE3-1] propagation requires both a functional J-domain and the tetratricopeptide repeat (TPR) domain, but surprisingly does not require Swa2-clathrin binding. Because the J-domain of Swa2 can be replaced with the J-domains of other proteins, our data strongly suggest that prion-chaperone specificity arises from the Swa2 TPR domain and supports a model where Swa2 acts through Hsp70, most likely to provide additional access points for Hsp104 to promote prion template generation. more...

Background: Analysis of process-related impurities is critical for the control of biopharmaceutical processes and the quality of final biological products. Residual impurities in monoclonal antibody products such as host cell proteins (HCPs) increase the risk of immunogenicity and may directly affect drug potency. Commonly used HCP ELISA often involves complicated sample preparation, lengthy operation, and large volumes of reagent. To overcome these challenges, a fully automated CHO HCP assay was developed using a microfluidic platform (MFP) system and compared with existing plate-based ELISA for quantification of HCP in monoclonal antibody purification intermediates. Results: The automated MFP based assay approach enabled an improved throughput (5–10-times faster), broader dynamic range (100-times) and decreased sample consumption, hands on time and duration for assay development compared with Tecan plate-based ELISA. Conclusion: The newly developed microfluidic assay demonstrated its advantages over pla... more...

The pathogenicity of different isolates of Fusarium oxysporum obtained from plants of Gerbera (Gerbera jamesonii), Chrysanthemum (Chrysanthemum morifolium), Paris daisy (Argyranthemum frutescens) and African daisy (Osteospermum sp.), all in the family Asteraceae, was tested on different cultivars of these hosts, to assess their pathogenicity. The reactions were compared with those of isolates of F. oxysporum f. sp. chrysanthemi and of f.sp. tracheiphilum obtained from the American Type Culture Collection. We found that isolates of F. oxysporum f. sp. chrysanthemi can be distinguished as three physiological races on the basis of their pathogenicity to the panel of differential cultivars. Sequencing of the intergenic spacer (IGS) region of ribosomal DNA (rDNA) and phylogenetic analysis showed that the Fusarium races fell into three phylogenetic groups, which coincided with those observed in pathogenicity tests. Analysis of the IGS sequences revealed a high degree of similarity among strains from Italy and Spain from different host species, suggesting that recent outbreaks in these ornamentals were probably caused by introduction of infected nursery material from a common origin. more...

CONDOTTA IN 11 DIVERSI COMUNI della provincia di Lecce, l’indagine era fi nalizzata a verifi - care l’incidenza di Xylella fastidiosa negli olivi con manifestazione conclamata di disseccamento rapido (OQDS), ovvero con chioma compromessa per oltre il 70%. I risultati hanno evidenziato la strettissima associazione tra il batterio e la malattia, a testimonianza dell’elevata sensibilità dei metodi diagnostici PCR ed ELISA in particolar modo nei focolai maturi (da almeno due anni). more...

This article has been accepted for publication in the Archives of Environmental Contamination and Toxicology. The final version can be accessed from the link below. This article has been made available through the Brunel Open Access Publishing Fund. Aquatic birds are commonly affected by oil spills. Despite rehabilitation efforts, the majority of rehabilitated common guillemots (Uria aalge) do not survive, whereas mute swans (Cygnus olor) tend to have higher post-release survival. Polyaromatic hydrocarbons (PAHs) present in crude oil and diesel are immunotoxic in birds affecting cell-mediated responses to immunogens. Because it is a target of PAH toxicity, T-lymphocyte response to controlled mitogen administration (phytohemagglutinnin test) was investigated in a scoping study as a potentially useful minimally invasive in vivo test of cell-mediated immunity. The test was performed on 69 mute swans and 31 common guillemots stranded on the Norfolk and Lincolnshire coastline and inland waterways in England (UK)either due to injury or to contamination with crude or diesel oil. T-lymphocyte response was significantly decreased in swans with greater oil scores. T-lymphocyte responses were also decreased in guillemots, but this finding was not statistically significant. more...

The random amplified polymorphic DNA (RAPD) technique was used to analyze the total genomic DNA of pathogenic isolates of Fusarium oxysporum on Gerbera jamesonii by comparing them to representatives of the formae speciales chrysanthemi and tracheiphilum. A close genetic relationship was observed among most of the new isolates from G. jamesonii. They shared RAPD markers with the tested representatives of the forma specialis chrysanthemi. Some isolates of those tested from diseased G. jamesonii were placed in a different cluster, which included representative isolates of forma specialis tracheiphilum. This is the first report of F. oxysporum f.sp. tracheiphilum on G. jamesonii. A rapid protocol for DNA extraction directly from fungal colonies grown on potato dextrose agar allowed complete analysis in less than 4 h. more...

This work examines diamond-like carbon (DLC) coatings deposited by plasma enhanced chemical vapour deposition (PECVD) as an environmentally friendly alternative to chromium plating in restoration of worn or damaged aircraft components. DLC coatings offer superior mechanical properties; however, high internal stresses and poor adhesion can prevent the deposition of thick films. This work examines a series of layered structures based on epoxy-resin interlayers with DLC applied as a surface film. Wear testing and examination with scanning electron microscopy and atomic force microscopy lead to the development of an optimum DLC/epoxy system with wear characteristics superior to those of chromium-plated steel. This new coating system has a great potential in restoring aircraft components in a more efficient and environmentally friendly manner. ?? 2009 The Surface Science Society of Japan. more...

Polycyclic aromatic hydrocarbons (PAHs) in crude oil cause a range of adverse effects in oiled seabirds, one of the most common being hemolytic anemia via oxidative attack of erythrocytes by PAH metabolites resulting in hemoglobin leakage and formation of Heinz bodies. In such cases, haptoglobin and ferritin are up-regulated to sequester free Hb and iron in the circulation. We investigated these plasma proteins as biomarkers of PAH-induced Heinz body hemolytic anemia in oiled seabirds. Concentration ranges of PAHs, HAP and FT in plasma samples were 10–184 ng/ml, 0–2.6 mg/ml and 0–7.6 ng/ml, respectively. Dose–response relationships between plasma PAH exposure and haptoglobin and ferritin (FT) were investigated, and evidence of erythrocyte Heinz body formation studied in 50 oiled common guillemots stranded on the Norfolk Wash coast (East England). Haptoglobin was negatively correlated, and FT was positively correlated with PAH exposure. Heinz bodies were also observed confirming the toxic mechanism causing hemolytic anemia and counts were positively correlated with exposure. Our results support the application of these complementary biomarkers to assess hemolytic effects of oiling in wildlife biomonitoring, which also discriminate the influence of hemolytic versus inflammatory effects in oiled guillemots. more...

Allergic disorders are characterized by Th2-polarization, thus physiological Th1-dependent mechanisms for fighting infectious diseases (ID) may be defective. This study aims at evaluating the number and duration of respiratory (RI) and gastrointestinal diseases (GI) in allergic and non-allergic subjects in a particular community: the crew of an operative warship consisting of 189 persons (171 males and 18 females). The study period lasted for 5 months during an operative mission. The number and duration (days) of RI and GI were evaluated. Allergic subjects experienced more numerous RI (p=0.0029) and more severe GI (p=0.0001) than non-allergic subjects. This preliminary study provides evidence that allergic subjects may have more numerous RI and prolonged GI than non-allergic subjects. more...

The literature pertaining to the use of registered antibacterial agents in Mediterranean finfish farming is reviewed, with an emphasis on the Greek fish-farming industry. This review provides a scientific resource dedicated to the design of future antibacterial dosing regimes in Mediterranean fish farming, where insufficient supporting information is currently available. This paper addresses the paucity in knowledge concerning pharmacokinetics and the efficacy and environmental impact of commonly used antibacterials needed to direct future research and promote good practices in the euryhaline fish farming industry. Several registered antibacterials are currently available for combating bacterial infections, including tetracyclines, (fluoro) quinolones, potentiated sulfa, penicillin and chloramphenicol derivatives. Based on the available data, oxytetracycline (OTC) and quinolone drugs (oxolinic acid – OA and flumequine – FLU) are the most widely used in Mediterranean aquaculture. As a result these drugs have received the most extensive studies, whereas, there is considerable paucity of reliable data on pharmacokinetic and the depletion characteristics of other drugs used, particularly potentiated sulfa, penicillin derivatives and florfenicol. We find there is incomplete data on drug efficacy and minimum inhibitory concentrations (MIC) for common antibacterials used against the major bacterial pathogens of Mediterranean fish species. Furthermore, a considerable lack of data on environmental drug concentrations around Mediterranean fish farms was also identified, highlighting the need for more extensive environmental studies to monitor contamination in environmental components i.e., water and sediment, and in non-target species (flora and fauna). Prudent selection and use of antibacterials can encourage lower dosage applications, enhance treatment efficacy, and help to minimize contamination of the environment. Selection of readily bioavailable drugs which have low environmental persistence, low aquatic toxicity and high antibacterial efficacy is advised, to reduce potential losses to the environment and associated toxic effects on target species and the development of bacterial resistance. Lack of present data made it impossible to provide thorough and accurate guidance on selection and use of antibacterials and approaches for minimizing environmental impacts for the treatment of major euryhaline aquaculture species. more...

The pharmacokinetics of oxolinic acid (OA) were investigated after a single intra-vascular injection (20 mg kg−1 fish) in sharpsnout sea bream (90 g), a promising new euryhaline species for Mediterranean fish farming. The distribution half-life (t1/2α) and the elimination half-life (t1/2β) of OA were calculated to be 0.4 and 10 h respectively. The apparent volume of distribution at steady-state (Vd(ss)) and total clearance rate (CLT) of the drug were found to be 2.1 L kg and 0.2 L kg−1 h−1 respectively. The bioavailability (F%) of OA following oral administration (40 mg kg−1 fish) was estimated to be 15%. The results indicate a rapid distribution and elimination of the drug, moderate tissue penetration, but low absorption in sharpsnout sea bream. The kinetic profile of OA found in this species is comparable with that observed in another well-known sparid, gilthead sea bream. more...

The pharmacokinetics of oxytetracycline (OTC) were investigated following single intravascular injection (40 mg/kg) in sharpsnout sea bream (90 g) at 19 °C. The distribution half-life (t1/2α) and the elimination half-life (t1/2β) of OTC were calculated to be 1.4 and 35 h, respectively. The apparent volume of distribution of the drug at steady-state (Vd(ss)) was found to be 4 l/kg. The total clearance rate (CLT) of the drug was low (0.08 l/kg h). Repeated attempts to investigate the bioavailability of OTC following oral administration (75 mg/kg; forced-fed or via the feed), revealed undetectable levels of the drug in plasma and muscle samples. Poor tissue absorption is likely due to significant hepatic metabolism of OTC in sharpsnout sea bream, but this requires further investigation. Thus, oral administration of OTC in farmed sharpsnout sea bream should be discouraged in favour of other routes of administration or other antibiotic drugs. more...

The tissue distribution and residue depletion of oxolinic acid (OA) were investigated in gilthead sea bream and sharpsnout sea bream after a multiple (10 days) in-feed administration (30 mg/kg fish) at 19 °C. Plasma, muscle, skin, liver and bile were sampled daily during treatment (1–10 days) and post-treatment (11,12, 14, 15 and 17 days) and subsequently analysed for OA concentrations using high-performance liquid chromatography and fluorescent detection (HPLC-SFD). Oxolinic acid displayed biphasic elimination in tissues of both species during treatment and reached “consumer safe levels” 24 h after treatment. OA concentration in gilthead sea bream peaked on day 4, with highest concentrations in liver (2660 ng/g), followed by bile (2370 ng/ml), skin (2350 ng/g), muscle (1330 ng/g) and plasma (880 ng/ml). In sharpsnout sea bream, peaked OA concentrations were observed in liver (2720 ng/g) on day 4, while highest concentrations were measured later on day 5 in plasma (910 ng/ml) and day 6 in bile (2360 ng/ml), skin (1960 ng/g) and muscle (1770 ng/g). The calculated areas under the concentration curves (AUC) also reflect a good tissue distribution of OA. Elimination half-lives (t1/2) were estimated to be 11–14 and 11–19 h in tissues of gilthead sea bream and sharpsnout sea bream, respectively. Multiple dosing of OA in sparids should be adequate to confront bacterial pathogens considering published minimum inhibitory concentrations of the drug. more...

This is the first study to investigate the pharmacokinetic parameters of oxytetracycline (OTC) in gilthead sea bream ( Sparus aurata ). Following a single intravascular injection of OTC (40 mg/kg) in 100 g fish kept at 20 °C, the distribution half-life ( t 1/2 α ) and the elimination half-life ( t 1/2 β ) of OTC were found to be long (2 and 53 h, respectively). The drug was well distributed throughout the internal body compartments since the apparent volume of distribution of the drug at steady-state ( V d(ss) ) and the apparent volume of distribution by the area method ( V d(area) ) were found to be high (2.9 and 3.8 l/kg, respectively). OTC had a long mean residence time (MRT) (56 h) but a short total clearance (CL T ) (0.05 l/kg h). The slow elimination of the drug suggests a need for long withdrawal periods prior to use of dosed fish for human consumption. Although the bioavailability ( F %) of OTC following oral administration in aqueous suspension (75 mg/kg) was calculated to be low (9%), maximum OTC plasma concentration was relatively high (2.5 μg/ml), indicating that current therapeutic regimens applied to gilthead sea bream might be effective against bacterial pathogens. more...

The concentrations of total mercury, polychlorinated biphenyls (PCBs), and organochlorine pesticides (SigmaDDT, dieldrin, hexachlorobenzene [HCB], and SigmaHCH) were determined in 61 whale meat products (bacon, blubber, red meat, liver, intestine, and tongue) purchased from retail outlets across Japan. Mean (range) concentrations of contaminants in all samples were: total mercury 4.17 (0.01-204); SigmaPCB 1.14 (0-8.94); SigmaDDT 0.98 (0-7.46); dieldrin 0.07 (0-0.35); HCB 0.06 (0-0.22); and SigmaHCH 0.07 (0-0.19) micro g/g (wet weight). The data were used to calculate estimated daily intakes (EDIs) of contaminants at two hypothetical levels of whale meat consumption. These EDIs were compared with FAO/WHO "tolerable daily intake" (TDI) values for each chemical. EDIs calculated for higher levels of whale meat consumption were in some cases exceptionally high and for many products exceeded FAO/ WHO-TDIs for total mercury, PCBs, and dieldrin, with exceedance factor values (EDI/TDI) for total mercury, PCBs, and dieldrin reaching maxima of 175, 5.36, and 2.1, respectively. For sensitive consumers and those with high-level consumption (e.g., whaling communities), exposure to mercury and to a lesser extent PCBs from certain whale blubber and bacon and striped dolphin liver products could lead to chronic health effects. The Japanese community should therefore exercise a precautionary approach to the consumption of such foods in excess, particularly by high-risk members of the population. more...

This is the first study on the pharmacokinetic parameters of oxolinic acid (OA) in gilthead sea bream, Sparus aurata L. The kinetic profile of OA was studied after a single intravascular injection (20 mg kg - 1 ) in 100 g fish at 20 °C. The distribution half-life (t 1 / 2 α ) and the elimination half-life (t 1 / 2 β ) of the drug were found to be short (0.51 and 12.60 h, respectively). The drug penetration from the plasma to the tissues was adequate as the apparent volume of distribution of the drug at steady-state (V d ( s s ) ) was found to be 2.11 L kg - 1 . The mean residence time (MRT) of OA was short (14.25 h) and the total clearance rate (Cl T ) of the drug was low (0.15 L kg - 1 h - 1 ). The bioavailability (F%) of OA following oral administration (30 mg kg - 1 ) was also low (14%). Maximum values were observed for muscle at 0.5 h after injection, with levels declining as with subsequent sampling. At the first two time points (0.5 and 1 h) plasma levels of OA were higher than muscle, however, the reverse was evident for subsequent samples. Following oral administration, highest muscle levels were found at 16 h and, with the exception of the 24-h sampling, muscle OA concentrations were higher than plasma at all time points. The fast elimination of OA suggests short withdrawal times with reference to human consumption of treated fish. more...

Yeast prions require a core set of chaperone proteins (Sis1, Ssa1, and Hsp104) for stable propagation in yeast, and yet some prions exhibit requirements for additional chaperone activities, indicat... more...

Polychlorinated biphenyl ( PCB) and dichlorodiphenylethane (DDE) methyl sulfone (MSF) metabolites possess high affinities for binding two homologous 16,000 Da homodimeric receptor proteins in the lung (Clara cell secretory protein, CCSP) and the uterus (uteroglobin, UG) , leading to selective bioaccumulation of MSFs in these tissues. As marine mammals are highly exposed to organochlorines, concentrations of PCBs, PCB MSFs, DDT, and DDE MSF were analyzed in blubber, lung, and uterus samples from harbor seal ( Phoca vitulina) and striped dolphin ( Stenella coeruleoalba) morbillivirus epizootic victims to investigate uterine and lung MSF accumulation. Mean uterus concentrations of PCB MSFs and DDE MSF in harbor seals were 0.61 and 0.04 μg/ g lipid weight and in striped dolphins 0.05 and 0.01 μg/ g lipid weight. Mean lung concentrations of PCB MSFs and DDE MSF in harbor seals were 0.96 and 0.02 μg/ g lipid weight and in striped dolphins 0.16 and 0.01 μg/ g lipid weight. To ascertain whether uterine and lung bioaccumulation of MSFs is possible due to the presence of CCSP and UG in seals, CCSP and UG proteins in uterine flushings and in uterine and lung and epithelial tissue from Baltic gray and ringed seals were characterized using gel electrophoresis and Western blotting techniques. UG- and CCSP-like proteins with molecular weights of 16,000 Da were resolved in all samples. This is the first demonstration of this protein in any marine mammalian species. The toxicological implications of MSF binding with UG and CCSP in marine mammals are discussed. Care for the Wild (UK) more...

Hepatic microsomal cytochrome P450, EROD and ECOD activity were investigated as biomarkers of PCB exposure in harbour seals (Phoca vitulina). Due to the difficulty of obtaining undegraded seal liver samples, standard spectrophotometric methodology was adapted to investigate P420 (degraded P450) as a PCB biomarker with partially degraded samples. Total PCB burdens in both blubber and liver had positive correlations with P450, P420 and MFO activity levels. The use of P420 biomarkers in this study supports the inclusion of samples from by-caught marine mammals for future biomonitoring studies. P450 isozymes CYP1A (P4501A) and CYP2B (P4502B) in conjunction with MFO activity were investigated as "specific" biomarkers of PCB exposure. They were found to reliably reflect levels of [MC] and [PB]-type PCB exposure in harbour seal liver. more...

Gerbera (Gerbera jamesonii) is one of the top 10 economically important flower crops in Europe as well as the United States. The acreage devoted to this crop continues to increase especially for use in landscape typologies. Abundant flowering from spring until autumn allows the use of this plant to decorate gardens, terraces, and borders. During the summer of 2009, an outbreak of a previously unknown powdery mildew was observed on potted gerbera ‘Mini Yellow’ growing in a private garden in Turin (northern Italy). Adaxial leaf surfaces were covered with white mycelium and conidia, and as the disease progressed, infected leaves turned yellow and died. Conidia were hyaline, ellipsoid, borne in chains (three conidia per chain), and measured 16 to 45 × 10 to 30 μm. Conidiophores measured 109 to 117 × 11 to 13 μm and had a foot cell measuring 72 to 80 × 11 to 12 μm followed by two shorter cells measuring 19 to 29 × 11 to 14 and 20 to 32 × 12 to 14 μm. Fibrosin bodies were absent and chasmothecia were not observed in the collected samples. On the basis of its morphology, the pathogen was identified as Golovinomyces cichoracearum. The internal transcribed spacer (ITS) region of rDNA was amplified with primers ITS1/ITS4 and sequenced. BLASTn analysis of the 548-bp fragment showed an E-value of 0.0 and a percentage homology of 99% with G. cichoracearum isolated from Coreopsis leavenworthii (Accession No. DQ871605) confirming diagnosis inferred by morphological analysis. The nucleotide sequence has been assigned GenBank Accession No. GQ870342. Pathogenicity was confirmed through inoculation by gently pressing diseased leaves onto leaves of three healthy potted plants of Gerbera ‘Mini Yellow’. Three noninoculated plants served as the control. Plants were maintained in a greenhouse at temperatures ranging between 20 and 30°C. Inoculated plants developed signs and symptoms after 8 days, whereas control plants remained healthy. The fungus present on inoculated plants was morphologically identical to that originally observed on diseased plants. To our knowledge, this is the first report of the presence of powdery mildew caused by G. cichoracearum on gerbera in Italy. Specimens are available at the Agroinnova Collection at the University of Torino. Gerbera is also susceptible to different powdery mildews. Powdery mildew of Gerbera jamesonii caused by Sphaerotheca fusca was reported in Italy (4). G. cichoracearum on Gerbera jamesonii was reported in North America (2), Argentina (3), and Switzerland (1). References: (1) A. Bolay. Cryptogam. Helv. 20:1, 2005. (2) M. Daughtrey et al. Page 39 in: Compendium of Flowering Potted Plant Diseases. The American Phytopathological Society, St Paul, MN, 1995. (3) R. Delhey et al. Schlechtendalia 10:79, 2003. (4) F. Zaccaria et al. Ann. Fac. Agrar. Univ. Stud. di Napoli Federico II 34:44, 2000. more...

A series of layered structures based on epoxy resins coated with diamond-like carbon (DLC) are examined as potential replacements for electroplated chromium in aerospace applications. Diamond-like carbon coatings can offer superior mechanical properties and tribological performance; however, in some applications high internal stresses and poor adhesion limit their practical use. A DLC/epoxy system is developed and studied utilising pin-on-disk testing, scanning electron microscopy and atomic force microscopy, resulting in an optimum system with characteristics superior to those of chromium-plated steel. This new coating system has a great potential in restoring worn or damaged aircraft components, without the health and environmental issues associated with chromium plating. The components can be rebuilt and improved over the original condition thus allowing an extension of service life and eliminating the need for costly replacements. more...

Aside from the physical effects of oiling (e.g., hypothermia, dehydration, emaciation), chronic toxicity of polycyclic aromatic hydrocarbons (PAHs) contamination is an important factor influencing long-term recovery of oiled sea birds following an oil spill. Monitoring PAH exposure can help identify populations at risk from toxic effects of PAHs for further study and/or protection. This is the first studyto quantify PAH and metabolite tissue burdens in sea birds directly oiled following oil spills. PAHs and hydroxylated PAHs were quantified in liver samples from oiled Common Guillemots (Uria aalge) stranded along the East Coast of England using gas chromatography-mass spectroscopy (GC-MS). Mean parent and metabolite PAH concentrations were 0.25+/-0.09 (range 0.04-0.97) and 0.52+/-0.14 (range 0.05-1.48) microg/g (wet wt.), respectively. The main source of PAH exposure was via ingestion of crude oil during preening, resulting in PAH uptake and tissue contamination beyond levels expected from exposure via the food chain. PAH composition corresponded with number of benzene rings in each compound and was typical of contamination from petrogenic sources; pentacyclictri- and tetracyclictricyclicdicyclic PAHs. The occurrence of PAH metabolites detected in liver samples also provided evidence of the presence and stereoselectivity of hepatic microsomal CYP1A1 in common guillemots. more...

External exposure assessment of oiled seabirds is undertaken by assessment of the percentage oil coverage of the plumage. Nondestructive monitoring of the toxic fraction of petroleum oils and diesels (polyaromatic hydrocarbons, PAHs) which enters the general circulation (internal exposure burden) of oiled seabirds is rarely undertaken. This is because the traditionally used chromatographic methods for plasma PAH analysis require larger sample volumes than those that can be safely collected from smaller species, such as guillemots (Uria aalge). Furthermore, these methods are not a cost-effective or practical approach for analysis of large numbers of birds in a short time period as part of an oil spill response in wildlife rehabilitation centers. This study describes the modification and validation of a commercially available PAH immunoassay (cRaPID PAH) to enable high-throughput, cost-effective, simple, and rapid determination of total PAH concentrations in 50 microL volumes of plasma. The limit of detection of the assay was 0.1 ng/mL as benzo-apyrene (BaP) equivalents with a working range of 0.120 ng/mL. As further validation of the immunoassay, PAHs were determined by GC-MS. GC-MS data were significantly positively correlated with corresponding immunoassay data for the same birds (r2 = 0.976, p < 0.001). The plasma PAH concentrations of 40 oiled guillemots stranded on U.K. shores were determined using the assay to demonstrate its usefulness for biomonitoring studies. The mean ,PAH concentration observed was 1.05 +/- 0.67 ppm (range 0.02-2.40 ppm as BaP equivalents). The modifications to the cRaPID PAH kit in this study enable nondestructive, high-throughput, semiquantitative determination of PAH concentrations in plasma samples suitable for exposure assessment of oiled seabirds during oil spill response and rehabilitation. more...

The potential for input of two common antibacterial agents in Mediterranean fish farms, oxytetracycline (OTC) and oxolinic acid (OA), was estimated from measurements of these drugs in the faecal excretions of two important farmed sparids, gilthead sea bream, Sparus aurata and sharpsnout sea bream Diplodus puntazzo. Oxolinic acid was found to be well absorbed by gilthead sea bream (92%) and sharpsnout sea bream (88%) while the absorption of OTC was found to be considerably lower in both species (27 and 40%, respectively). These data were integrated with production records for sparids, drug dosage regimes and treatment frequency information to calculate potential annual drug release to the aquatic environment from Greek fish farms. These calculations suggest potentially significant quantities of unmetabolised OTC can be passed unabsorbed through the body of treated sparids and excreted via the faeces into the local marine environment. The situation with OA was much less pronounced. It was estimated that potentially more than 1900 kg of OTC and more than 50 kg of OA may be released via faecal excretion into the environment by sparid farms per year. Further drug may also be released via uneaten medicated feed, leached drugs and other routes of fish elimination (renal excretion, branchial secretions). Drug pollution of the marine environment in the vicinity of fish farms can have adverse ecological effects, including development of resistant bacterial populations and exposure with potential drug accumulation in aquatic fauna and flora. more...

The present study investigated the kinetic profile of flumequine (FLU) in gilthead sea bream Sparus aurata (170 g) held at 19 degrees C and evaluated its in vitro efficacy against important bacterial diseases in Mediterranean mariculture. Following a single intravascular injection (10 mg kg(-1) fish), the distribution half-life (t1/2alpha) and the half-life of the terminal phase of elimination (t1/2gamma) of the drug were 0.2 and 30 h respectively. Tissue penetration of FLU was low, since both the apparent distribution volume of the drug at steady-state (Vd(SS)) and the apparent volume of the central compartment (Vc) were small (0.57 and 0.15 l kg(-1)). The mean residence time (MRT) was short (11 h) and the total clearance (CL(T)) of the drug was slow (0.05 l kg(-1) h(-1)). Following oral administration (20 mg kg(-1)), the bioavailability (F %) of FLU was 29% and the maximum plasma concentration was 1.7 microg ml(-1). The minimum inhibitory concentration (MIC) of the drug in distilled water supplemented with 2% NaCl against Vibrio anguillarum Serotype 1b, Photobacterium damsela ssp. piscicida, V. alginolyticus, V. damsela and V. fluvialis was 0.15, 0.3, 1.2, 0.019 and 0.15 microg ml(-1) respectively. The addition however of 10 mM Ca2+ and 55 mM Mg2+ to the medium resulted in an 8- to >120-fold reduction in FLU activity. The results indicate that FLU has an adequate kinetic profile in gilthead sea bream and that marine cations induce a significant impact on the activity of FLU, rendering its use against bacterial pathogens questionable. more...

Hepatic cytochrome P-450 (CYP450) isozymes are involved in xenobiotic detoxification and steroid metabolism. Seals are highly exposed to polychlorinated biphenyls (PCBs) in the environment, resulting in CYP450 induction, which may have concomitant effects on CYP450 steroid metabolism. Experiments were conducted to determine rates of steroid metabolism in harbor seal (Phoca vitulina) liver. CYP450 was induced with increasing liver PCB concentrations between 0.2 and 5.73 microg/g (wet weight). Progesterone metabolism was significantly decreased with increasing liver PCB concentration. Testosterone metabolism was also increased with increasing liver PCB concentration. The results indicate a change in steroid metabolism associated with increasing PCB burden, suggesting that PCBs may have a modulating effect on hepatic steroid biotransformation in environmentally exposed seals. The possible implications of this for endocrine homeostasis in seals are discussed. more...

The levels of PCBs and PCB methyl sulphone metabolites were determined in the blubber of six species of cetaceans that originated from the Irish Sea and the Aegean Sea (Mediterranean). Burdens ranging from 2.80 to 27.80 microg g-1 (lipid weight) total PCBs and 0.03 to 0.58 microg g-1 (lipid weight) total methyl sulphones were recorded. The highest cetacean methyl sulphone burden (0.58 microg g-1) was in an Irish Sea harbor porpoise (Phocoena phocoena). Methyl sulphone isomer formation capacity, as determined by PCB:MSF ratios, varied considerably between species as follows: harbour porpoise (1:10)pilot whale (Globicephalus melas), white-sided dolphin (Lagenorhyncus acutus) (1:50)common dolphin (Delphinus delphis), Risso's dolphin (Grampus griseus), and striped dolphin (Stenella coerueoalba), (1:100), and was related to interspecific variation in PCB methyl sulphone formation capacity. more...

0044-8486/$ - see front matter D 2003 Elsevier B.V. All rights reserved.doi:10.1016/j.aquaculture.2004.06.029DOI of original article 10.1016/j.aquaculture.2003.10.016.* Corresponding author. Laboratory of Fish Nutrition and Pathology, National Centre for Marine Research,Aghios Kosmas 16604, Ellinikon, Attiki, Greece. Tel.: +30 210 9829238; fax: +30 210 1 9811713.E-mail address: grigos@ncmr.gr (G. Rigos).Aquaculture 240 (2004) 629www.elsevier.com/locate/aqua-online more...

In this paper we study, in the framework of Sobolev weight spaces, differential operators of elliptic type, with singular coefficients, perturbed by a potential. Conditions for compactness of embedding Ws,σr,p(Ω)↪Ws,00,p(Ω)=Lsp(Ω) for density ofD(Ω) on Ws,σr,p(Ω) and discretness conditions for the spectrum of Aσ in the Banach spaces Lsp(Ω), have been established. more...