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Volume 3, Problème 2 (2009)

article de recherche

Improved and simplified recombineering approach for influenza virus reverse genetics

Qinfang Liu, Shuai Wang, Guangpeng Ma, Juan Pu, Nicole E Forbes, Earl G Brown and Jin-Hua Liu

Typical reverse genetics systems for generating influenza viruses require the insertion of each genome segments by DNA ligation into vectors for genome synthesis and expression. Herein is described the construction and use of a novel pair of plasmid vectors for cloning all eight genome segments of influenza A virus by homologous recombination for influenza virus reverse genetics. Plasmids, pLLBA and pLLBG, were constructed to possess opposing RNA polymerase I and RNA polymerase II transcription units for generating influenza genomic and messenger RNAs, respectively. In addition these promoters flanked a recombination cassette which comprised the conserved 5’ (13bp) and 3’ (12bp) terminal promoters of influenza virus. These vectors differed due to the presence of an A or a G (plus sense) to correspond to differences at nucleotide position 4 among negative-sense influenza virus promoters. The cloning approach involved homologous recombination of each influenza gene segment and the appropriate linearized pLLBA or pLLBG vectors in E. coli. Direct cloning by recombination was simpler and faster than conventional restriction digestion and ligation methods. This new vector system was successfully used to clone and rescue various influenza viruses and thus has the potential to promote the rapid analysis and vaccine development of novel influenza strains.

Communication courte

Heterosubtypic anti-avian H5N1 influenza antibodies in intravenous immunoglobulins from globally separate populations protect against H5N1 infection in cell culture

John S Sullivan1,2, Paul W Selleck3, Teena Downton, Ingrid Boehm, Anna-Maree Axel, Yasmin Ayob, Natalie M Kapitza, Wayne Dyer, Anna Fitzgerald, Bradley Walsh and Garry W Lynch

With antigenically novel epidemic and pandemic influenza strains persistently on the horizon it is of fundamental importance that we understand whether heterosubtypic antibodies gained from exposures to circulating human influenzas exist and can protect against emerging novel strains. Our studies of IVIG obtained from an infection-naive population (Australian) enabled us to reveal heterosubtypic influenza antibodies that cross react with H5N1. We now expand those findings for an Australian donor population to include IVIG formulations from a variety of northern hemisphere populations. Examination of IVIGs from European and South East-Asian (Malaysian) blood donor populations further reveal heterosubtypic antibodies to H5N1 in humans from different global regions. Importantly these protect against highly pathogenic avian H5N1 infection in vitro, albeit at low titres of inhibition. Although there were qualitative and quantitative differences in binding and protection between globally different formulations, the heterosubtypic antibody activities for the respective IVIGs were in general quite similar. Of particular note because of the relative geographic proximity to the epicentre of H5N1 and the majority of human infections, was the similarity in the antibody binding responses between IVIGs from the Malayan peninsula, Europe and Australia. These findings highlight the value of employing IVIGs for the study of herd immunity, and particularly heterosubtypic antibody responses to viral antigens such as those conserved between circulating human influenzas and emerging influenza strains such as H5N1. They also open a window into a somewhat ill defined arena of antibody immunity, namely heterosubtypic immunity.

Article court

Genetic variance of Derzsy's disease strains isolated in Poland

Grzegorz Woźniakowski, Wojciech Kozdruń and Elżbieta Samorek-Salamonowicz

The aim of this study was the assessment of the genetic variance of Derzy’s disease (GPV) strains isolated from cases occurring in Poland. The nucleotide and predicted aminoacid sequences of VP2 and VP3 surface proteins of the Polish GPV strains were compared with other strains previously isolated in Hungary, France, Germany, China and Taiwan. The observed genetic variance of the aminoacid sequence within the group of Polish strains was low and reached 5% of the overall analysed sequence. Considerable differences in aminoacid sequence were found in the case of Polish field GPV strains and Muscovy duck parvovirus strain MDPV FM which was also analysed in this study. The conducted investigations confirmed the presupposition that Polish GPV strains and strains previously isolated in Hungary and France share a common origin.

Article court

Acquired heterosubtypic antibodies in human immunity for avian H5N1 influenza

Garry W Lynch, Paul Selleck and John S Sullivan

Well understood are the adaptive and dramatic neutralizing homosubtypic antibody responses to hypervariable, immunodominant sites of the hemagglutinin (HA) and neuraminidase (NA) of individual influenza strains. These define influenza subtypes and vaccines modelled upon their HA and NA antigens provide seasonal neutralizing antibody protection against subsequent exposure to the strain and its close relatives, but give little if any protection against antigenically drifted or shifted strains. Contrasting to this is a different form of acquired antibody response, called heterosubtypic immunity. This provides a more seasoned adaptive antibody response to immunerecessive epitopes that are highly-conserved amongst strains. Although, such responses are of lower individual amplitudes than seasonal mechanisms they are active across influenza subtypes, and may give pre-emptive protection against new strains yet to emerge. Heterosubtypic immunities have been well studied in animals, but surprisingly there is minimal evidence for this type of antibody immunity in humans. Thus championed is the notion that seasoned humoral responses can through repeated exposure to sites widely conserved across different strains, cumulatively provide humans with a level of broad protection against emergent novel strains, such as H5N1, that is not afforded by seasonal humoral responses.

Article court

Avian influenza vaccination in Egypt: Limitations of the current strategy

Marisa Peyre, Hamid Samaha, Yilma Jobre Makonnen, Ahmed Saad, Amira Abd-Elnabi, Saber Galal, Toni Ettel, Gwenaelle Dauphin, Juan Lubroth, Fran?ois Roger, Joseph Domenech

Vaccination of domestic poultry against avian influenza (AI) has been used on a large-scale in South East Asia since 2003 and in Egypt since 2006 to fight H5N1 highly-pathogenic avian influenza (HPAI) epidemics. The decision to use mass vaccination against HPAI in Egypt was taken as an emergency measure based on positive impact of such control measures in Vietnam and the People’s Republic of China. However, three years on, the impact on disease control of AI vaccination in Egypt has been very limited. Despite the continuous vaccination of poultry against HPAI, poultry outbreaks and human cases are reported regularly. A recent assessment study highlighted substantial weaknesses in the current immunisation programme and its lack of positive impact on the spread of infection or the maintenance of public health safety. The shortcomings of the vaccination strategy may be attributed in part to a lack of sufficient support in terms of funding and communication, the absence of an efficient monitoring system, and inadequate training of field technicians. The difficulties of blanket vaccinations in semi-commercial farms and household poultry sectors are well known, however, improvements in the industrial sector should be possible though better government controls and greater collaboration with the private sector. AI vaccination should be regarded as just one control tool within a broader disease control program integrating surveillance, outbreak investigation, disease management systems, and the rigorous implementation of bio-security measures. If incorrectly implemented, AI vaccination has a limited impact as a disease control measure. Moreover, without strict bio-security precautions undertaken during its application, farm visits to vaccinate poultry could facilitate the spread of the virus and therefore become a risk factor with important implications on the maintenance of the virus and potential risk for human exposure.

Article de perspective

Origins of the Spanish Influenza pandemic (1918-1920) and its relation to the First World War

Anton Erkoreka

The virus which was responsible for the first benign wave of the Spanish Influenza in the spring of 1918, and which was to become extremely virulent by the end of the summer of 1918, was inextricably associated with the soldiers who fought during the First World War. The millions of young men who occupied the military camps and trenches were the substrate on which the influenza virus developed and expanded. Many factors contributed to it, such as: the mixing on French soil of soldiers and workers from the five continents, the very poor quality of life of the soldiers, agglomeration, stress, fear, war gasses used for the first time in history in a massive and indiscriminate manner, life exposed to the elements, cold weather, humidity and contact with birds, pigs and other animals, both wild and domestic. Today, this combination of circumstances is not present and so it seems unlikely that new pandemics, such as those associated with the avian influenza or swine influenza, will emerge with the virulence which characterized the Spanish Influenza during the autumn of 1918.

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