Elke Lange

Friedrich Loeffler Institute, Greifswald, Mecklenburg-Vorpommern, Germany

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Publications (47)132.27 Total impact

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    ABSTRACT: In this study we describe the use of a rope-in-a-bait sampling method ("pSWAB": pathogen sampling wild animals with baits) for non-invasive saliva sampling aimed at the detection of foot-and-mouth disease (FMD) viral genome in wild boar. The pSWABs are produced in the form of a standardized product by embedding a 10cm long cotton rope in a cereal-based bait matrix. To assess the general suitability of this novel sampling technique an animal experiment was conducted to detect FMD viral genome in saliva of infected wild boar. Two juvenile animals were inoculated in the bulb of the heel with a recent wild boar FMD virus isolate and kept together with three noninoculated wild boar of the same age. Over a period of 29 days, the animals were sampled by using five pSWABs per day in addition to the collection of blood and conventional saliva swabs taken every three to four days. Viral RNA in pSWABs was identified already 24h after infection during the incubation period and until 23dpi. Comparison of the results of pSWAB sampling with those of conventional saliva swabs or serum samples showed satisfactory sensitivity. These experimental data demonstrate the suitability of non-invasive sampling of wild boar by using pSWABs as a sensitive, cheap and feasible sample collection technique independent of hunting activities. In addition, the use of non-invasive sampling in an appropriate surveillance strategy is discussed.
    Veterinary microbiology. 05/2014;
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    ABSTRACT: Pigs can be severely harmed by flu, and represent important reservoir hosts, in which new human pathogens like the recent pandemic swine-origin H1N1 influenza A virus can arise by mutation and reassortment of genome segments. To obtain novel, safe influenza vaccines for pigs, and to investigate the antigen-specific immune response, we modified an established live-virus vaccine against Aujeszkys disease of swine, pseudorabies virus (PrV) strain Bartha, to serve as vector for the expression of hemagglutinin (HA) of swine-origin H1N1 virus. To facilitate transgene insertion, the genome of PrV-Ba was cloned as a bacterial artificial chromosome. HA expression occurred under control of the human or murine cytomegalovirus immediate early promoters (P-HCMV, P-MCMV), but could be substantially enhanced by synthetic introns and adaptation of the codon usage to that of PrV. However, despite abundant expression, the heterologous glycoprotein was not detectably incorporated into mature PrV particles. Replication of HA-expressing PrV in cell culture was only slightly affected compared to that of the parental virus strain. A single immunization of pigs with the PrV vector expressing the codon-optimized HA gene under control of P-MCMV induced high levels of HA-specific antibodies. The vaccinated animals were protected from clinical signs after challenge with a related swine-origin H1N1 influenza A virus, and challenge virus shedding was significantly reduced.
    Journal of General Virology 01/2014; · 3.13 Impact Factor
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    ABSTRACT: Swine influenza viruses (SIV) regularly cause significant disease in pigs worldwide. Since there is no causative treatment of SIV, we tested if probiotic Enterococcus (E.) faecium NCIMB 10415 or zinc (Zn) oxide as feed supplements provide beneficial effects upon SIV infection in piglets. Seventy-two weaned piglets were fed three different diets containing either E. faecium or different levels of Zn (2500 ppm, Zn(high); 50 ppm, Zn(low)). Half of the piglets were vaccinated intramuscularly (VAC) twice with an inactivated trivalent SIV vaccine, while all piglets were then infected intranasally with H3N2 SIV. Significantly higher weekly weight gains were observed in the E. faecium group before virus infection, and piglets in Zn(high) and E. faecium groups gained weight after infection while those in the control group (Zn(low)) lost weight. Using ELISA, we found significantly higher H3N2-specific antibody levels in the E. faecium+VAC group 2 days before and at the day of challenge infection as well as at 4 and 6 days after challenge infection. Higher hemagglutination inhibition (HI) titers were also observed in the Zn(high)+VAC and E. faecium+VAC groups at 0, 1 and 4 days after infection. However, there were no significant differences in virus shedding and lung lesions between the dietary groups. Using flow cytometry analysis significantly higher activated T helper cells and cytotoxic T lymphocyte percentages in the PBMCs were detected in the Zn(high) and E. faecium groups at single time points after infection compared to the Zn(low) control group, but no prolonged effect was found. In the BAL cells no influence of dietary supplementation on immune cell percentages could be detected. Our results suggest that feeding high doses of zinc oxide and particularly E. faecium could beneficially influence humoral immune responses after vaccination and recovery from SIV infection, but not affect virus shedding and lung pathology.
    PLoS ONE 01/2014; 9(1):e87007. · 3.53 Impact Factor
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    ABSTRACT: Serological investigations of swine influenza virus infections and epidemiological conclusions thereof are challenging due to the complex and regionally variable pattern of co-circulating viral subtypes and lineages and varying vaccination regimes. Detection of subtype-specific antibodies currently depends on hemagglutination inhibition (HI) assays which are difficult to standardize and unsuitable for large scale investigations. The nucleocapsid protein (NP) and HA1 fragments of the hemagglutinin protein (HA) of five different lineages (H1N1av, H1N1pdm, H1pdmN2, H1N2, H3N2) of swine influenza viruses were bacterially expressed and used as diagnostic antigens in indirect ELISA. Proteins were co-translationally mono-biotinylated and refolded in vitro into an antigenically authentic conformation. Western blotting and indirect ELISA revealed highly subtype-specific antigenic characteristics of the recombinant HA1 proteins although some cross reactivity especially among antigens of the H1 subtype were evident. Discrimination of antibodies directed against four swine influenza virus subtypes co-circulating in Germany was feasible using the indirect ELISA format. Bacterially expressed recombinant NP and HA1 swine influenza virus proteins served as antigens in indirect ELISAs and provided an alternative to commercial blocking NP ELISA and HI assays concerning generic (NP-specific) and HA subtype-specific sero-diagnostics, respectively, on a herd basis.
    Virology Journal 07/2013; 10(1):246. · 2.09 Impact Factor
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    ABSTRACT: The emergence of the human 2009 pandemic H1N1 (H1N1pdm) from swine populations re-focused public and scientific attention towards swine as an important source of influenza A viruses bearing zoonotic potential. Widespread and year-round circulation of at least four stable lineages of porcine influenza viruses in Germany in a region with a high density swine population between 2009 and 2012 is documented here. European avian-derived H1N1 (H1N1av) viruses dominated the epidemiology, followed by human-derived subtypes H1N2 and H3N2. H1N1pdm viruses and, in particular, recently emerged reassortants between H1N1pdm and porcine HxN2 viruses (H1pdmN2) were detected in about 8% of cases. Further reassortants between these main lineages were diagnosed sporadically. On-going diversification both at the phylogenetic and the antigenic level was evident for the H1N1av lineage and for some of its reassortants. The H1avN2 reassortant R1931/11 displayed conspicuously distinct genetic and antigenic features and easily transmitted from pig-to-pig in an experimental infection. Continuing diverging evolution was also observed in the H1pdmN2 lineage. These viruses carry seven genome segments of the H1N1pdm virus including a hemagglutinin gene which encodes an antigenically markedly altered protein. The zoonotic potential of this lineage remains to be determined. The results highlight the relevance of surveillance and control of porcine influenza virus infections. This is important with respect to health status of swine herds. In addition, a more exhaustive tracing of the formation, transmission and spread of new reassortant influenza A viruses with unknown zoonotic potential is urgently required.
    Journal of Virology 07/2013; · 5.08 Impact Factor
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    ABSTRACT: Swine influenza virus (SIV) is not only an important respiratory pathogen in pigs but also a threat to human health. The pandemic influenza A(H1N1)pdm09 virus likely originated in swine through reassortment between a North American triple reassortant and Eurasian avian-like SIV. The North American triple reassortant virus has genes from avian, human and swine influenza viruses. An effective vaccine may protect the pork industry from economic losses and curb the development of new virus variants that may threaten public health. In the present study, we evaluated the efficacy of a recombinant equine herpesvirus type 1 (EHV-1) vaccine (rH_H1) expressing the hemagglutinin H1 of A(H1N1)pdm09 in the natural host. Our data shows that the engineered rH_H1 vaccine induces influenza virus-specific antibody responses in pigs and is able to protect at least partially against challenge infection: no clinical signs of disease were detected and virus replication was reduced as evidenced by decreased nasal virus shedding and faster virus clearance. Taken together, our results indicate that recombinant EHV-1 encoding H1 of A(H1N1)pdm09 may be a promising alternative for protection of pigs against infection with A(H1N1)pdm09 or other influenza viruses.
    Virus Research 01/2013; · 2.75 Impact Factor
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    ABSTRACT: Despite substantial improvements, influenza vaccine production-and availability-remain suboptimal. Influenza vaccines based on mRNA may offer a solution as sequence-matched, clinical-grade material could be produced reliably and rapidly in a scalable process, allowing quick response to the emergence of pandemic strains. Here we show that mRNA vaccines induce balanced, long-lived and protective immunity to influenza A virus infections in even very young and very old mice and that the vaccine remains protective upon thermal stress. This vaccine format elicits B and T cell-dependent protection and targets multiple antigens, including the highly conserved viral nucleoprotein, indicating its usefulness as a cross-protective vaccine. In ferrets and pigs, mRNA vaccines induce immunological correlates of protection and protective effects similar to those of a licensed influenza vaccine in pigs. Thus, mRNA vaccines could address substantial medical need in the area of influenza prophylaxis and the broader realm of anti-infective vaccinology.
    Nature Biotechnology 11/2012; · 32.44 Impact Factor
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    ABSTRACT: In domestic pigs strict control measures and the use of gene-deleted marker vaccines resulted in the elimination of pseudorabies virus (PrV) infections in many parts of Europe and North America. In free-roaming feral pigs and wild boar populations, however, serological surveys and monitoring in The Americas, Europe and North Africa provided serological and virological evidence that PrV is more widely distributed than previously assumed. Thus, there is a constant risk of spillover of PrV infection from wild pig populations to domestic animals which could require intervention to limit the infection in wild pigs. To investigate whether oral immunization of wild boar by live-attenuated PrV could be an option, wild boar and domestic pigs were orally immunized with 2×10(6) TCID(50) of the attenuated live PrV vaccine strain Bartha supplied either with a syringe or within a blister, and subsequently intranasally challenged with 10(6) TCID(50) of the highly virulent PrV strain NIA-3. Oral immunization with live-attenuated PrV was able to confer protection against clinical signs in wild boar and against transmission of challenge virus to naïve contact animals. Only two vaccinated domestic pigs developed neurological signs after challenge infection. Our results demonstrate that oral immunization against PrV infection in wild boar is possible. In case increasing PrV infection rates in wild boar may enhance the risk for spillover into domestic pig populations, oral immunization of wild boar against PrV in endemic areas might be a feasible control strategy.
    Veterinary Microbiology 07/2012; · 3.13 Impact Factor
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    ABSTRACT: The incursion of the human pandemic influenza A virus H1N1 (2009) (H1N1 pdm) into pig populations and its ongoing co-circulation with endemic swine influenza viruses (SIVs) has yielded distinct human-porcine reassortant virus lineages. The haemagglutinin (HA) gene of H1N1 pdm was detected in 41 influenza virus-positive samples from seven swine herds in north-west Germany in 2011. Eight of these samples yielded virus that carried SIV-derived neuraminidase N2 of three different porcine lineages in an H1N1 pdm backbone. The HA sequences of these viruses clustered in two distinct groups and were distinguishable from human and other porcine H1 pdm by a unique set of eight non-synonymous mutations. In contrast to the human population, where H1N1 pdm replaced seasonal H1N1, this virus seems to co-circulate and interact more intensely with endemic SIV lineages, giving rise to reassortants with as-yet-unknown biological properties and undetermined risks for public health.
    Journal of General Virology 05/2012; 93(Pt 8):1658-63. · 3.13 Impact Factor
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    ABSTRACT: Due to the tremendous socio-economic impact of classical swine fever (CSF) outbreaks, emergency vaccination scenarios are continuously under discussion. Unfortunately, all currently available vaccines show restrictions either in terms of marker capacities or immunogenicity. Recent research efforts were therefore directed at the design of new modified live marker vaccines. Among the most promising candidates the chimeric pestiviruses "CP7_E2alf" and "flc11" were identified. Within an international research project, these candidates were comparatively tested in challenge experiments after a single oral vaccination. Challenge infection was carried out with highly virulent CSF virus strain "Koslov", 14 or 21 days post vaccination (dpv), respectively. Safety, efficacy, and marker potential were addressed. All assessments were done in comparison with the conventional "gold standard" C-strain "Riems" vaccine. In addition to the challenge trials, multiple vaccinations with both candidates were performed to further assess their marker vaccine potential. All vaccines were safe and yielded full protection upon challenge 21 days post vaccination. Neither serological nor virological investigations showed major differences among the three vaccines. Whereas CP7_E2alf also provided clinical protection upon challenge at 14 days post vaccination, only 50% of animals vaccinated with flc11, and 83% vaccinated with C-strain "Riems" survived challenge at this time point. No marked differences were seen in protected animals. Despite the fact that all multiple-vaccinated animals stayed sero-negative in the accompanying marker test, the discriminatory assay remains a weak point due to delayed or inexistent detection of some of the vaccinated and subsequently infected animals. Nevertheless, the potential as live marker vaccines could be confirmed for both vaccine candidates. Future efforts will therefore be directed at the licensing of "Cp7_E2alf" as the first live marker vaccine for CSF.
    Veterinary Microbiology 02/2012; 158(1-2):42-59. · 3.13 Impact Factor
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    ABSTRACT: Porcine reproductive and respiratory syndrome (PRRS) causes economic losses in the pig industry worldwide, and PRRS viruses (PRRSV) are classified into the two distinct genotypes "North American (NA, type 2)" and "European (EU, type 1)". In 2006, a highly pathogenic NA strain of PRRSV (HP-PRRSV), characterized by high fever as well as high morbidity and mortality, emerged in swine farms in China. Therefore, a real-time reverse transcription polymerase chain reaction (RT-qPCR) assay specific for HP-PRRSV was developed and combined with type 1- and type 2-specific RT-qPCR systems. Furthermore, an internal control, based on a heterologous RNA, was successfully introduced. This final multiplex PRRSV RT-qPCR, detecting and typing PRRSV, had an analytical sensitivity of less than 200 copies per µl for the type 1-assay and 20 copies per µl for the type 2- and HP assays and a high diagnostic sensitivity. A panel of reference strains and field isolates was reliably detected and samples from an animal trial with a Chinese HP-PRRS strain were used for test validation. The new multiplex PRRSV RT-qPCR system allows for the first time the highly sensitive detection and rapid differentiation of PRRSV of both genotypes as well as the direct detection of HP-PRRSV.
    PLoS ONE 01/2012; 7(6):e38251. · 3.53 Impact Factor
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    ABSTRACT: Despite considerable host species barriers, interspecies transmissions of influenza A viruses between wild birds, poultry and pigs have been demonstrated repeatedly. In particular, viruses of the subtypes H1 and H3 were transmitted between pigs and poultry, predominantly turkeys, in regions with a high population density of both species. The recovery of a swine influenza H1N1 virus from a turkey flock in Germany in 2009 prompted us to investigate molecularly the subtype H1 viruses recently detected in wild birds, pigs and poultry. The goal of this study was to investigate the relationship between H1N1 viruses originating from wild and domestic animals of Germany and to identify potential trans-species transmission or reassortment events. Hemagglutinin and neuraminidase gene or full-length genome sequences were generated from selected, current H1N1 viruses from wild birds, pigs and turkeys. Phylogenetic analyses were combined with genotyping and analyses of the deduced amino acid sequences with respect to biologically active sites. Antigenic relationships were assessed by hemagglutination inhibition reactions. Phylogenetic analysis of the hemagglutinin sequences showed that viruses from distinct H1 subgroups co-circulate among domestic animals and wild birds. In addition, these viruses comprised different genotypes and were distinguishable antigenically. An H1N1 virus isolated from a turkey farm in northern Germany in 2009 showed the highest similarity with the avian-like porcine H1N1 influenza viruses circulating in Europe since the late 1970s. The data demonstrate the genetic and antigenic heterogeneity of H1 viruses currently circulating in domestic and wild animals in Germany and points to turkeys as a possible bridge between avian and mammalian hosts.
    Influenza and Other Respiratory Viruses 07/2011; 5(4):276-84. · 1.47 Impact Factor
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    ABSTRACT: A natural reassortant influenza A virus consisting of seven genome segments from pandemic (H1N1) 2009 virus and a neuraminidase segment from a Eurasian porcine H1N1 influenza A virus was detected in a pig herd in Germany. The obvious reassortment compatibility between the pandemic (H1N1) 2009 and H1N1 viruses of porcine origin raises concern as to whether swine may become a reservoir for further reassortants of pandemic (H1N1) 2009 viruses with unknown implications for human health and swine production.
    Journal of General Virology 02/2011; 92(Pt 5):1184-8. · 3.13 Impact Factor
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    ABSTRACT: To determine susceptibility of chickens, turkeys, and mice to pandemic (H1N1) 2009 virus, we conducted contact exposure and inoculation experiments. We demonstrated that chickens were refractory to infection. However, oculo-oronasally inoculated turkeys and intranasally inoculated mice seroconverted without clinical signs of infection.
    Emerging Infectious Diseases 04/2010; 16(4):703-5. · 6.79 Impact Factor
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    ABSTRACT: Natural infections with different subtypes of low pathogenicity avian influenza viruses (LPAIVs) are very common in wild duck populations. Recent outbreaks of high pathogenicity avian influenza virus (HPAIV) H5N1 in Eurasian and African countries stimulated monitoring activities in aquatic wild bird populations. Surveillance mainly focused on virus detection. Only a few serologic investigations have been conducted so far, although such data may retrospectively elucidate epidemiologic patterns of different AIV subtypes in the populations under study. To better understand the immunologic and serologic reactions of mallards after infection with LPAIV, we investigated the AIV type- and subtype-specific antibody dynamics in mallards after different LPAIV infections by hemagglutination inhibition, competitive enzyme-linked immunosorbent assay, and western blot analysis, as appropriate. Four groups of mallard ducks were used: 1) naturally infected birds, 2) birds that were experimentally infected with LPAIV, 3) birds that were immunized with inactivated virus preparations, and 4) negative control birds. Ducks were monitored for up to 15 mo, and serum samples were investigated every 1-4 wk. It could be shown that infection with LPAIV in mallards can be traced serologically over prolonged periods of time.
    Avian Diseases 03/2010; 54(1):79-85. · 1.73 Impact Factor
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    ABSTRACT: SummaryA horse imported from Brazil developed a respiratory illness 2 weeks after arrival in Germany. After an initial but inefficient treatment glanders was diagnosed based on serological and molecular biological findings. The present case highlights the potential risk of an importation of glanders in free areas. The fact that veterinarians in countries where glanders has been eradicated for decades are not familiar with the clinical symptoms of the disease, can favour the entry of the disease. In order to prevent the spread of glanders, the sanctions of the veterinary authorities in such cases of the infection are of utmost importance.
    Equine Veterinary Education. 01/2010; 21(3):147 - 150.
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    ABSTRACT: The RIEMSER Schweinepestoralvakzine is an attenuated vaccine for oral vaccination of wild boar against classical swine fever (CSF). The safety of this licensed bait vaccine which is based on the CSF virus (CSFV) strain "C" was investigated in eight animal species, e.g. weaner pigs (n=111), wild boar (n=11), ruminants (cattle, goats and sheep, n=11), foxes (n=5), rabbits (n=12), and mice (n=10). Animals were vaccinated either with a single vaccine dose containing at least 10(4.5) TCID(50), or with overdoses, i.e. the 10-fold dose, or they were subjected to repeated application schemes. During the entire observation period none of the animals which were given the vaccine virus showed clinical signs, with the exception of rabbits. These reacted to the vaccination with fever. Orally vaccinated pigs did not transmit vaccine virus to susceptible contact animals (sentinels). In none of the species examined neither vaccine virus nor viral RNA could be detected in blood after vaccination. In one wild boar viral RNA could be established in the tonsil 21 days post-vaccination (dpv); all other organ samples tested virologically negative. Up to 77.5% of the pigs and wild boar developed virus neutralising antibodies (VNA) already 14 dpv. The mean VNA titres observed in the vaccination groups seemed to depend rather on individual factors than on the administered virus dose (virus titre per dose) or the vaccination scheme. These results are comparable with findings obtained during oral vaccination campaigns in wild boar and after parenteral vaccination with this C-strain virus. From the results presented here it can be concluded that RIEMSER Schweinepestoralvakzine is safe for target and non-target species.
    Veterinary Microbiology 11/2009; 143(2-4):133-8. · 3.13 Impact Factor
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    ABSTRACT: A new chimeric pestivirus "CP7_E1E2alf_TLA", based on the infectious cDNA of bovine viral diarrhea virus (BVDV) strain CP7, was constructed. The substitution of BVDV E1 and E2 with the respective proteins of classical swine fever (CSF) strain Alfort 187 allows an optimal heterodimerization of E1 and E2 in the chimeric virus, which is beneficial for efficient and authentic virus assembly and growth. In addition, for implementation of E2-based marker diagnostics, the previously described antigenic CSFV-specific TAVSPTTLR epitope was exchanged with the corresponding E2-epitope of BVDV strain CP7. Recombinant virus CP7_E1E2alf_TLA displayed a growth defect, and was not reacting with monoclonal antibodies used in commercial E2 antibody blocking ELISAs. Therefore, efficacy as well as marker properties of CP7_E1E2alf_TLA were investigated in an animal experiment with both a high dose and a low dose vaccine preparation. All CP7_E1E2alf_TLA-vaccinated animals seroconverted until day 28 post-vaccination with neutralizing antibodies. Furthermore, at the day of challenge infection CP7_E1E2alf_TLA-immunized animals showed distinct lower ELISA values in a commercial CSFV E2 antibody test in comparison to the C-strain vaccinated controls. However, E2-ELISA reactivity as well as neutralizing titers were directly connected to the dosage used for vaccination, and only the low dose group had E2-ELISA values below threshold until challenge infection. Following challenge infection with highly virulent CSFV strain Koslov, all vaccinees were protected, however, short-term fever episodes and very limited CSFV genome detection with very low copy numbers could be observed. In conclusion, manipulation of the TAVSPTTLR-epitope within the tested chimeric virus resulted in an slightly reduced efficacy, but the E2 marker properties unexpectedly did not allow a clear differentiation of infected from vaccinated animals in some cases.
    Veterinary Microbiology 10/2009; 142(1-2):45-50. · 3.13 Impact Factor
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    ABSTRACT: Due to the vast economic consequences of classical swine fever (CSF) outbreaks, emergency vaccination plans are under discussion in European Union Member States. However, animals vaccinated with the conventional C-strain vaccine are subject to trade restrictions. To ease these restrictions, potent marker vaccines are required. One promising candidate is the chimeric pestivirus CP7_E2alf. For emergency vaccination in a CSF outbreak scenario, early onset of immunity is required. Here, the studies performed with a CP7_E2alf virus stock produced under good manufacturing conditions (GMP) are reported. In challenge experiments, CP7_E2alf induced full clinical protection 1 week after intramuscular vaccination, and 2 weeks after oral immunization. Furthermore, even after application of diluted vaccine preparations complete protection could be achieved if challenge infection was carried out 4 weeks after vaccination. In conclusion, GMP-produced CP7_E2alf proved to be a suitable marker vaccine candidate - also for emergency vaccination - both after intramuscular and oral application.
    Vaccine 10/2009; 27(47):6522-9. · 3.77 Impact Factor
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    ABSTRACT: Influenza virus A/H1N1, which is currently causing a pandemic, contains gene segments with ancestors in the North American and Eurasian swine lineages. To get insights into virus replication dynamics, clinical symptoms and virus transmission in pigs, we infected animals intranasally with influenza virus A/Regensburg/D6/09/H1N1. Virus excretion in the inoculated pigs was detected in nasal swabs from 1 day post-infection (p.i.) onwards and the pigs developed generally mild symptoms, including fever, sneezing, nasal discharge and diarrhoea. Contact pigs became infected, shed virus and developed clinical symptoms similar to those in the inoculated animals. Plasma samples of all animals remained negative for virus RNA. Nucleoprotein- and haemagglutinin H1-specific antibodies could be detected by ELISA 7 days p.i. CD4(+) T cells became activated immediately after infection and both CD4(+) and CD8(+) T-cell populations expanded from 3 to 7 days p.i., coinciding with clinical signs. Contact chickens remained uninfected, as judged by the absence of virus excretion, clinical signs and seroconversion.
    Journal of General Virology 08/2009; 90(Pt 9):2119-23. · 3.13 Impact Factor