NS1 proteins of avian influenza A viruses can act as antagonists of the human alpha/beta interferon response

Many viruses, including human influenza A virus, have developed strategies for counteracting the host type I interferon (IFN) response. We have explored whether avian influenza viruses were less capable of combating the type I IFN response in mammalian cells, as this might be a determinant of host range restriction. A panel of avian influenza viruses isolated between 1927 and 1997 was assembled. The selected viruses showed variation in their ability to activate the expression of a reporter gene under the control of the IFN-beta promoter and in the levels of IFN induced in mammalian cells. Surprisingly, the avian NS1 proteins expressed alone or in the genetic background of a human influenza virus controlled IFN-beta induction in a manner similar to the NS1 protein of human strains. There was no direct correlation between the IFN-beta induction and replication of avian influenza viruses in human A549 cells. Nevertheless, human cells deficient in the type I IFN system showed enhanced replication of the avian viruses studied, implying that the human type I IFN response limits avian influenza viruses and can contribute to host range restriction.

Studies on particulate matter (PM10) and its precursors over urban environment of Reading, UK

The extent to which airborne particles penetrate into the human respiratory system is determined mainly by their size, with possible health effects. The research over the scientific evidence of the role of airborne particles in adverse health effects has been intensified in recent years. In the present study, seasonal variations of PM10 and its relation with anthropogenic activities have been studied by using the data from UK National Air Quality Archive over Reading, UK. The diurnal variation of PM10 shows a morning peak during 7:00-10:00 LT and an evening peak during 19:00-22:00 LT. 3 The variation between 12:00 and 17:00 LT remains more or less steady for PM10 with the minimum value of similar to 16 mu g m(-3). PM10 and black smoke (BS) concentrations during weekdays were found to be high compared to weekends. A reduction in the concentration of PM10 has been found during the Christmas holidays compared to normal days during December. Seasonal variations of PM10 showed high values during spring compared to other seasons. A linear relationship has been found between PM10 and NO, during March, July, November and December suggesting that most of the PM10 is due to local traffic exhaust emissions. PM10 and SO2 concentrations showed positive correlation with the correlation coefficient of R-2 = 0.65 over the study area. Seasonal variations of SO2 and NOx showed high concentrations during winter and low concentrations during spring. Fraction of BS in PM10 has been found to be 50% during 2004 over the study area. (C) 2005 Elsevier Ltd. All rights reserved.

Deposition of charged inhaled aerosols with transient airflow in sequential lung airway model

Transport and deposition of charged inhaled aerosols in double planar bifurcation representing generation three to five of human respiratory system has been studied under a light activity breathing condition. Both steady and oscillatory laminar inhalation airflow is considered. Particle trajectories are calculated using a Lagrangian reference frame, which is dominated by the fluid force driven by airflow, gravity force and electrostatic forces (both of space and image charge forces). The particle-mesh method is selected to calculate the space charge force. This numerical study investigates the deposition efficiency in the three-dimensional model under various particle sizes, charge values, and inlet particle distribution. Numerical results indicate that particles carrying an adequate level of charge can improve deposition efficiency in the airway model.

A single amino acid in the HA of pH1N1 2009 influenza virus affects cell tropism in human airway epithelium, but not transmission in ferrets

The first pandemic of the 21(st) century, pandemic H1N1 2009 (pH1N1 2009), emerged from a swine-origin source. Although human infections with swine-origin influenza have been reported previously, none went on to cause a pandemic or indeed any sustained human transmission. In previous pandemics, specific residues in the receptor binding site of the haemagglutinin (HA) protein of influenza have been associated with the ability of the virus to transmit between humans. In the present study we investigated the effect of residue 227 in HA on cell tropism and transmission of pH1N1 2009. In pH1N1 2009 and recent seasonal H1N1 viruses this residue is glutamic acid, whereas in swine influenza it is alanine. Using human airway epithelium, we show a differential cell tropism of pH1N1 2009 compared to pH1N1 2009 E227A and swine influenza suggesting this residue may alter the sialic acid conformer binding preference of the HA. Furthermore, both pH1N1 2009 E227A and swine influenza multi-cycle viral growth was found to be attenuated in comparison to pH1N1 2009 in human airway epithelium. However this altered tropism and viral growth in human airway epithelium did not abrogate respiratory droplet transmission of pH1N1 2009 E227A in ferrets. Thus, acquisition of E at residue 227 was not solely responsible for the ability of pH1N1 2009 to transmit between humans.

Mutations in H5N1 influenza virus hemagglutinin that confer binding to human tracheal airway epithelium

The emergence in 2009 of a swine-origin H1N1 influenza virus as the first pandemic of the 21st Century is a timely reminder of the international public health impact of influenza viruses, even those associated with mild disease. The widespread distribution of highly pathogenic H5N1 influenza virus in the avian population has spawned concern that it may give rise to a human influenza pandemic. The mortality rate associated with occasional human infection by H5N1 virus approximates 60%, suggesting that an H5N1 pandemic would be devastating to global health and economy. To date, the H5N1 virus has not acquired the propensity to transmit efficiently between humans. The reasons behind this are unclear, especially given the high mutation rate associated with influenza virus replication. Here we used a panel of recombinant H5 hemagglutinin (HA) variants to demonstrate the potential for H5 HA to bind human airway epithelium, the predominant target tissue for influenza virus infection and spread. While parental H5 HA exhibited limited binding to human tracheal epithelium, introduction of selected mutations converted the binding profile to that of a current human influenza strain HA. Strikingly, these amino-acid changes required multiple simultaneous mutations in the genomes of naturally occurring H5 isolates. Moreover, H5 HAs bearing intermediate sequences failed to bind airway tissues and likely represent mutations that are an evolutionary "dead end." We conclude that, although genetic changes that adapt H5 to human airways can be demonstrated, they may not readily arise during natural virus replication. This genetic barrier limits the likelihood that current H5 viruses will originate a human pandemic.

Mapping the immune response to the outer domain of a human immunodeficiency virus-1 clade C gp120

The outer domain (OD) of human immunodeficiency virus (HIV)-1 gp120 represents an attractive, if difficult, target for a beneficial immune response to HIV infection. Unlike the entire gp120, the OD is structurally stable and contains the surfaces that interact with both the primary and secondary cellular receptors. The primary strain-specific neutralizing target, the V3 loop, lies within the OD, as do epitopes for two cross-reactive neutralizing monoclonal antibodies (mAbs), b12 and 2G12, and the contact sites for a number of inhibitory lectins. The OD is poorly immunogenic, at least in the context of complete gp120, but purposeful OD immunization can lead to a substantial antibody response. Here, we map the antibody generated following immunization with a clade C OD. In contrast to published data for the clade B OD, the majority of the polyclonal response to the complete clade C OD is to the V3 loop; deletion of the loop substantially reduces immunogenicity. When the loop sequence was substituted for the epitope for 2F5, a well-characterized human cross-neutralizing mAb, a polyclonal response to the epitope was generated. A panel of mAbs against the clade C OD identified two mAbs that reacted with the loop and were neutralizing for clade C but not B isolates. Other mAbs recognized both linear and conformational epitopes in the OD. We conclude that, as for complete gp120, V3 immunodominance is a property of OD immunogens, that the responses can be neutralizing and that it could be exploited for the presentation of other epitopes.

Generation of candidate human influenza vaccine strains in cell culture: rehearsing the European response to an H7N1 pandemic threat

Background: Although H5N1 avian influenza viruses pose the most obvious imminent pandemic threat, there have been several recent zoonotic incidents involving transmission of H7 viruses to humans. Vaccines are the primary public health defense against pandemics, but reliance on embryonated chickens eggs to propagate vaccine and logistic problems posed by the use of new technology may slow our ability to respond rapidly in a pandemic situation. Objectives: We sought to generate an H7 candidate vaccine virus suitable for administration to humans whose generation and amplification avoided the use of eggs. Methods: We generated a suitable H7 vaccine virus by reverse genetics. This virus, known as RD3, comprises the internal genes of A/Puerto Rico/8/34 with surface antigens of the highly pathogenic avian strain A/Chicken/Italy/13474/99 (H7N1). The multi-basic amino acid site in the HA gene, associated with high pathogenicity in chickens, was removed. Results: The HA modification did not alter the antigenicity of the virus and the resultant single basic motif was stably retained following several passages in Vero and PER. C6 cells. RD3 was attenuated for growth in embryonated eggs, chickens, and ferrets. RD3 induced an antibody response in infected animals reactive against both the homologous virus and other H7 influenza viruses associated with recent infection by H7 viruses in humans. Conclusions: This is the first report of a candidate H7 vaccine virus for use in humans generated by reverse genetics and propagated entirely in mammalian tissue culture. The vaccine has potential use against a wide range of H7 strains.

Infection of human airway epithelium by human and avian strains of influenza a virus

We describe the characterization of influenza A virus infection of an established in vitro model of human pseudostratified mucociliary airway epithelium (HAE). Sialic acid receptors for both human and avian viruses, alpha-2,6- and alpha-2,3-linked sialic acids, respectively, were detected on the HAE cell surface, and their distribution accurately reflected that in human tracheobronchial tissue. Nonciliated cells present a higher proportion of alpha-2,6-linked sialic acid, while ciliated cells possess both sialic acid linkages. Although we found that human influenza viruses infected both ciliated and nonciliated cell types in the first round of infection, recent human H3N2 viruses infected a higher proportion of nonciliated cells in HAE than a 1968 pandemic-era human virus, which infected proportionally more ciliated cells. In contrast, avian influenza viruses exclusively infected ciliated cells. Although a broad-range neuraminidase abolished infection of HAE by human parainfluenza virus type 3, this treatment did not significantly affect infection by influenza viruses. All human viruses replicated efficiently in HAE, leading to accumulation of nascent virus released from the apical surface between 6 and 24 h postinfection with a low multiplicity of infection. Avian influenza A viruses also infected HAE, but spread was limited compared to that of human viruses. The nonciliated cell tropism of recent human H3N2 viruses reflects a preference for the sialic acid linkages displayed on these cell types and suggests a drift in the receptor binding phenotype of the H3 hemagglutinin protein as it evolves in humans away from its avian virus precursor.

Restrictions to the adaptation of influenza A virus H5 hemagglutinin to the human host

The binding specificities of a panel of avian influenza virus subtype H5 hemagglutinin (RA) proteins bearing mutations at key residues in the receptor binding site were investigated. The results demonstrate that two simultaneous mutations in the receptor binding site resulted in H5 RA binding in a pattern similar to that shown by human viruses. Coexpression of the ion channel protein, M2, from most avian and human strains tested protected H5 RA conformation during trafficking, indicating that no genetic barrier to the reassortment of the H5 surface antigen gene with internal genes of human viruses existed at this level.

Interactions of decay-accelerating factor (DAF) with haemagglutinating human enteroviruses: utilizing variation in primate DAF to map virus binding sites

A cellular receptor for the haemagglutinating enteroviruses (HEV), and the protein that mediates haemagglutination, is the membrane complement regulatory protein decay accelerating factor (DAF; CD55). Although primate DAF is highly conserved, significant differences exist to enable cell lines derived from primates to be utilized for the characterization of the DAF binding phenotype of human enteroviruses. Thus, several distinct DAF-binding phenotypes of a selection of HEVs (viz. coxsackievirus A21 and echoviruses 6, 7, 11-13, 29) were identified from binding and infection assays using a panel of primate cells derived from human, orang-utan, African Green monkey and baboon tissues. These studies complement our recent determination of the crystal structure of SCR(34) of human DAF [Williams, P., Chaudhry, Y., Goodfellow, I. G., Billington, J., Powell, R., Spiller, O. B., Evans, D. J. & Lea, S. (2003). J Biol Chem 278, 10691-10696] and have enabled us to better map the regions of DAF with which enteroviruses interact and, in certain cases, predict specific virus-receptor contacts.

Control of human immunodeficiency virus type-1 protease activity in insect cells expressing Gag-Pol rescues assembly of immature but not mature virus-like particles

Expression of human immunodeficiency virus type 1 (HIV-1) Gag protein in insect cells using baculovirus vectors leads to the abundant production of virus-like particles (VLPs) that represent the immature form of the virus. When Gag-Pol is included, however, VLP production is abolished, a result attributed to premature protease activation degrading the intracellular pool of Gag precursor before particle assembly can occur. As large-scale synthesis of mature noninfectious VLPs would be useful, we have sought to control HIV protease activity in insect cells to give a balance of Gag and Gag-Pol that is compatible with mature particle formation. We show here that intermediate levels of protease activity in insect cells can be attained through site-directed mutagenesis of the protease and through antiprotease drug treatment. However, despite Gag cleavage patterns that mimicked those seen in mammalian cells, VLP synthesis exhibited an essentially all-or-none response in which VLP synthesis occurred but was immature or failed completely. Our data are consistent with a requirement for specific cellular factors in addition to the correct ratio of Gag and Gag-Pol for assembly of mature retrovirus particles in heterologous cell types. (C) 2003 Elsevier Science (USA). All rights reserved.

Receptor binding profiles of avian influenza virus hemagglutinin subtypes on human cells as a predictor of pandemic potential

The host adaptation of influenza virus is partly dependent on the sialic acid (SA) isoform bound by the viral hemagglutinin (HA). Avian influenza viruses preferentially bind the α-2,3 SA and human influenza viruses the α-2,6 isoform. Each isoform is predominantly associated with different surface epithelial cell types of the human upper airway. Using recombinant HAs and human tracheal airway epithelial cells in vitro and ex vivo, we show that many avian HA subtypes do not adhere to this canonical view of SA specificity. The propensity of avian viruses to adapt to human receptors may thus be more widespread than previously supposed.

Human enhancement: could you become infected with a computer virus?

Experiments demonstrating human enhancement through the implantation of technology in healthy humans have been performed for over a decade by some academic research groups. More recently, technology enthusiasts have begun to realize the potential of implantable technology such as glass capsule RFID transponders. In this paper it is argued that implantable RFID devices have evolved to the point whereby we should consider the devices themselves as simple computers. Presented here is the infection with a computer virus of an RFID device implanted in a human. Coupled with our developing concept of what constitutes the human body and its boundaries, it is argued that this study has given rise to the world’s first human infected with a computer virus. It has taken the wider academic community some time to agree that meaningful discourse on the topic of implantable technology is of value. As developments in medical technologies point to greater possibilities for enhancement, this shift in thinking is not too soon in coming.

Characterization of virus-like particles associated with the human faecal and caecal microbiota

This work represents an investigation into the presence, abundance and diversity of virus-like particles (VLPs) associated with human faecal and caecal samples. Various methodologies for the recovery of VLPs from faeces were tested and optimized, including successful down-stream processing of such samples for the purpose of an in-depth electron microscopic analysis, pulsed-field gel electrophoresis and efficient DNA recovery. The applicability of the developed VLP characterization method beyond the use of faecal samples was then verified using samples obtained from human caecal fluid.

Broadly neutralizing antibodies protect against hepatitis C virus quasispecies challenge

A major problem in hepatitis C virus (HCV) immunotherapy or vaccine design is the extreme variability of the virus. We identified human monoclonal antibodies (mAbs) that neutralize genetically diverse HCV isolates and protect against heterologous HCV quasispecies challenge in a human liver-chimeric mouse model. The results provide evidence that broadly neutralizing antibodies to HCV protect against heterologous viral infection and suggest that a prophylactic vaccine against HCV may be achievable.