Aurélie Sausy

Prevalence and seasonality of tick-borne pathogens in questing Ixodes ricinus ticks from Luxembourg.

ABSTRACT In Europe, ixodid ticks are important arthropod vectors of human and animal pathogens, but comprehensive studies of the prevalence of all relevant pathogens in Central Europe are scarce. As a result of ecological changes, the incidences of tick-borne infections are expected to increase. In this study, 1,394 nymphal and adult Ixodes ricinus ticks sampled monthly during the active season from 33 ecologically distinct collection sites throughout Luxembourg were screened for all human tick-borne pathogens relevant in Central Europe. Species were identified by sequence analysis of detection PCR amplicons. Mean infection rates of ticks were 11.3% for Borrelia burgdorferi sensu lato, 5.1% for Rickettsia sp., 2.7% for Babesia sp., and 1.9% for Anaplasma phagocytophilum. No tick was found to be infected with Coxiella sp., Francisella tularensis subsp., or Tick-borne encephalitis virus (TBEV). A total of 3.2% of ticks were infected with more than one pathogen species, including mixed Borrelia infections (1.5%). Seasonal variations of tick infection rates were observed for Borrelia, Babesia, and Anaplasma, possibly reflecting a behavioral adaptation strategy of questing ticks. A positive correlation between the grade of urbanization and Borrelia infection rate of ticks was observed, suggesting an established urban zoonotic cycle. We also found Hepatozoon canis (0.1%) and Bartonella henselae (0.3%), which so far have not been found in questing Ixodes ricinus ticks in Central Europe.

Exceptional genetic variability of hepatitis B virus indicates that Rwanda is east of an emerging African genotype E/A1 divide

In Western Africa, hepatitis B virus (HBV) genotype E predominates throughout a vast crescent spanning from Senegal to Namibia and at least to the Central African Republic to the East. Although from most of the eastern parts of sub‐Saharan Africa only limited sets of strains have been characterized, these belong predominantly to genotype A. To study how far the genotype E crescent extends to the East, a larger number of HBV strains from Rwanda were analyzed. Phylogenetic analysis of 45 S fragment sequences revealed strains of genotypes A (n = 30), D (n = 10), C (n = 4), and B (n = 1). Twelve genotype A sequences formed a new cluster clearly separated from the reference strains of the known sub‐genotypes. Thus, with four genotypes and at least six sub‐genotypes and a new cluster of genotype A strains, HBV shows an exceptional genetic variability in this small country, unprecedented in sub‐Saharan Africa. Despite this exceptional genetic variability, not a single genotype E virus was found indicating that this country does not belong to the genotype E crescent, but is east of an emerging African genotype E/A1 divide. J. Med. Virol. 81:435–440, 2009.

Swine Influenza Virus Antibodies in Humans, Western Europe, 2009

Serologic studies for swine influenza viruses (SIVs) in humans with occupational exposure to swine have been reported from the Americas but not from Europe. We compared levels of neutralizing antibodies against 3 influenza viruses—pandemic (H1N1) 2009, an avian-like enzootic subtype H1N1 SIV, and a 2007–08 seasonal subtype H1N1—in 211 persons with swine contact and 224 matched controls in Luxembourg. Persons whose profession involved contact with swine had more neutralizing antibodies against SIV and pandemic (H1N1) 2009 virus than did the controls. Controls also had antibodies against these viruses although exposure to them was unlikely. Antibodies against SIV and pandemic (H1N1) 2009 virus correlated with each other but not with seasonal subtype H1N1 virus. Sequential exposure to variants of seasonal influenza (H1N1) viruses may have increased chances for serologic cross-reactivity with antigenically distinct viruses. Further studies are needed to determine the extent to which serologic responses correlate with infection.

Tannins from Hamamelis virginiana Bark Extract: Characterization and Improvement of the Antiviral Efficacy against Influenza A Virus and Human Papillomavirus

Antiviral activity has been demonstrated for different tannin-rich plant extracts. Since tannins of different classes and molecular weights are often found together in plant extracts and may differ in their antiviral activity, we have compared the effect against influenza A virus (IAV) of Hamamelis virginiana L. bark extract, fractions enriched in tannins of different molecular weights and individual tannins of defined structures, including pseudotannins. We demonstrate antiviral activity of the bark extract against different IAV strains, including the recently emerged H7N9, and show for the first time that a tannin-rich extract inhibits human papillomavirus (HPV) type 16 infection. As the best performing antiviral candidate, we identified a highly potent fraction against both IAV and HPV, enriched in high molecular weight condensed tannins by ultrafiltration, a simple, reproducible and easily upscalable method. This ultrafiltration concentrate and the bark extract inhibited early and, to a minor extent, later steps in the IAV life cycle and tannin-dependently inhibited HPV attachment. We observed interesting mechanistic differences between tannin structures: High molecular weight tannin containing extracts and tannic acid (1702 g/mol) inhibited both IAV receptor binding and neuraminidase activity. In contrast, low molecular weight compounds (<500 g/mol) such as gallic acid, epigallocatechin gallate or hamamelitannin inhibited neuraminidase but not hemagglutination. Average molecular weight of the compounds seemed to positively correlate with receptor binding (but not neuraminidase) inhibition. In general, neuraminidase inhibition seemed to contribute little to the antiviral activity. Importantly, antiviral use of the ultrafiltration fraction enriched in high molecular weight condensed tannins and, to a lesser extent, the unfractionated bark extract was preferable over individual isolated compounds. These results are of interest for developing and improving plant-based antivirals.

Characterization of Newcastle Disease Viruses in Wild and Domestic Birds in Luxembourg from 2006 to 2008

ABSTRACT Newcastle disease virus (NDV) is one of the most important viral diseases of birds. Wild birds constitute a natural reservoir of low-virulence viruses, while poultry are the main reservoir of virulent strains. Exchange of virus between these reservoirs represents a risk for both bird populations. Samples from wild and domestic birds collected between 2006 and 2010 in Luxembourg were analyzed for NDV. Three similar avirulent genotype I strains were found in ducks during consecutive years, suggesting that the virus may have survived and spread locally. However, separate introductions cannot be excluded, because no recent complete F gene sequences of genotype I from other European countries are available. Detection of vaccine-like strains in wild waterbirds suggested the spread of vaccine strains, despite the nonvaccination policy in Luxembourg. Among domestic birds, only one chicken was positive for a genotype II strain differing from the LaSota vaccine and exhibiting a so-far-unrecognized fusion protein cleavage site of predicted low virulence. Three genotype VI strains from pigeons were the only virulent strains found. The circulation of NDV in wild and free-ranging domestic birds warrants continuous surveillance because of increased concern that low-virulence wild-bird viruses could become more virulent in domestic populations.

Identification of a CD4 T-cell epitope in the hemagglutinin stalk domain of pandemic H1N1 influenza virus and its antigen-driven TCR usage signature in BALB/c mice

The stalk region of the influenza virus hemagglutinin is relatively well conserved compared with the globular head domain, which makes it a potential target for use as a universal vaccine against influenza. However, the role of CD4 T cells in the hemagglutinin stalk-specific immune response is not clear. Here we identified a mouse CD4 T-cell epitope that encompasses residues HA2113-131 from the hemagglutinin stalk domain after a sub-lethal infection of influenza. In response to stimulation with the identified epitope, splenocytes derived from the infected mice showed significant polyfunctionality as shown by IL-2, TNF-α and IFN-γ production as well as degranulation. Moreover, mice immunized with the peptide corresponding to this CD4 T-cell epitope exhibited interindividual sharing of the CD4 T-cell receptor β sequences, and they had a higher survival rate following a challenge with a lethal dose of pandemic H1N1 influenza virus. Thus, our data demonstrated a crucial role of hemagglutinin stalk-specific CD4 T cells in the host immune response against influenza virus infection.

Serological evidence of pandemic (H1N1) 2009 virus in pigs, West and Central Africa

Besides birds, pigs are another important reservoir of influenza A viruses that can be transmitted to human, as highlighted by the emergence and spread of the pandemic (H1N1) virus (pdm/09) in 2009. Surveillance in pigs is therefore necessary for public health and influenza pandemic preparedness. Nevertheless, there is a serious lack of data on influenza in Africa, especially in swine. We therefore collected serum samples from pigs in Nigeria (2009, 2012) and Cameroon (2011) in which the presence of anti-influenza A neutralizing antibodies was investigated. Our serological survey suggests that, before the 2009 pandemic, only rare swine and human H3N2 or human H1N1 infections occurred in Nigeria in swine. However, in 2011-2012, 27.4% of pigs in Nigeria and 5.6% in Cameroon had antibodies against H1N1 viruses. Higher antibody titres against pdm/09 suggested that pigs were exposed to this or a similar virus, either by multiple introductions or sustained circulation, and that reactivity against American and European swine H1N1 viruses resulted from cross-reaction.

Genetic Diversity of the Hepatitis B Virus Strains in Cuba: Absence of West-African Genotypes despite the Transatlantic Slave Trade

Cuba is an HBsAg low-prevalence country with a high coverage of anti-hepatitis B vaccine. Its population is essentially the result of the population mix of Spanish descendants and former African slaves. Information about genetic characteristics of hepatitis B virus (HBV) strains circulating in the country is scarce. The HBV genotypes/subgenotypes, serotypes, mixed infections, and S gene mutations of 172 Cuban HBsAg and HBV-DNA positive patients were determined by direct sequencing and phylogenetic analysis. Phylogenetic analysis of HBV S gene sequences showed a predominance of genotype A (92.4%), subgenotype A2 (84.9%) and A1 (7.6%). Genotype D (7.0%) and subgenotype C1 (0.6%) were also detected but typical (sub)genotypes of contemporary West-Africa (E, A3) were conspicuously absent. All genotype A, D, and C strains exhibited sequence characteristics of the adw2, ayw2, and adrq serotypes, respectively. Thirty-three (19.1%) patients showed single, double, or multiple point mutations inside the Major Hydrophilic domain associated with vaccine escape; eighteen (10.5%) patients had mutations in the T-cell epitope (amino acids 28-51), and there were another 111 point mutations downstream of the S gene. One patient had an HBV A1/A2 mixed infection. This first genetic study of Cuban HBV viruses revealed only strains that were interspersed with strains from particularly Europe, America, and Asia. The absence of genotype E supports previous hypotheses about an only recent introduction of this genotype into the general population in Africa. The presence of well-known vaccine escape (3.5%) and viral resistance mutants (2.9%) warrants strain surveillance to guide vaccination and treatment strategies.

Novel Alphacoronaviruses and Paramyxoviruses co-circulate with Type 1 and SARS-related Betacoronaviruses in synanthropic bats in Luxembourg.

ABSTRACT Several infectious disease outbreaks with high mortality in humans have been attributed to viruses that are thought to have evolved from bat viruses. In this study from Luxembourg, the genetic diversity and epidemiology of paramyxoviruses and coronaviruses shed by the bat species Rhinolophus ferrumequinum and Myotis emarginatus were evaluated. Feces collection (n = 624) was performed longitudinally in a mixed-species colony in 2015 and 2016. In addition, feces (n = 254) were collected cross-sectionally from six Myotis emarginatus colonies in 2016. By use of degenerate primers in a nested format, overall prevalences of 1.1% (10/878) and 4.9% (43/878) were determined for paramyxoviruses and coronaviruses. Sequences of the partial RNA-dependent RNA polymerase and spike glycoprotein genes of coronaviruses, as well as sequences of the partial L gene of paramyxoviruses, were obtained. Novel paramyxovirus and Alphacoronavirus strains were identified in different Myotis emarginatus colonies, and severe acute respiratory syndrome (SARS)-related Betacoronavirus strains were shed by Rhinolophus ferrumequinum. Logistic regression revealed that the level of Alphacoronavirus shedding was highest in July (odds ratio, 2.8; P < 0.01), probably due to periparturient stress. Phylogenetic analyses point to close virus-host coevolution, and the high genetic similarity of the study strains suggests that the Myotis emarginatus colonies in Luxembourg are socially connected. Most interestingly, we show that bats also host Betacoronavirus1 strains. The high similarity of the spike gene sequences of these viruses with mammalian Betacoronavirus 1 strains may be of concern. Both the SARS-related and Betacoronavirus 1 strains detected in bats in Luxembourg may cross the species barrier after a host adaptation process. IMPORTANCE Bats are a natural reservoir of a number of zoonotic pathogens. Several severe outbreaks in humans (e.g., a Nipah virus outbreak in Malaysia in 1998, and the almost global spread of severe acute respiratory syndrome in 2003) have been caused by bat-borne viruses that were transmitted to humans mostly after virus adaptation (e.g., in intermediate animal hosts). Despite the indigenousness of bat species that host viruses with suspected zoonotic potential and despite the zoonotic transmission of European bat 1 lyssavirus in Luxembourg, knowledge about the diversity and epidemiology of bat viruses remains limited in this country. Moreover, in contrast to other European countries, bat viruses are currently not included in the national surveillance activities of this land-locked country. We suggest that this gap in disease surveillance should be addressed, since we show here that synanthropic bats host viruses that may be able to cross the species barrier.

Proteome profiling of virus-host interactions of wild type and attenuated measles virus strains.

UNLABELLED Quantitative gel-based proteomics (2D DIGE coupled to MALDI-TOF/TOF MS) has been used to investigate the effects of different measles virus (MV) strains on the host cell proteome. A549/hSLAM cells were infected either with wild type MV strains, an attenuated vaccine or a multiple passaged Vero cell adapted strain. By including interferon beta treatment as a control it was possible to distinguish between the classical antiviral response and changes induced specifically by the different strains. Of 38 differentially expressed proteins in total (p-value ≤0.05, fold change ≥2), 18 proteins were uniquely modulated following MV infection with up to 9 proteins specific per individual strain. Interestingly, wt strains displayed distinct protein patterns particularly during the late phase of infection. Proteins were grouped into cytoskeleton, metabolism, transcription/translation, immune response and mitochondrial proteins. Bioinformatics analysis revealed mostly changes in proteins regulating cell death and apoptosis. Surprisingly, wt strains affected the cytokeratin system much stronger than the vaccine strain. To our knowledge, this is the first study on the MV-host proteome addressing interstrain differences. BIOLOGICAL SIGNIFICANCE In the present study we investigated the host cell proteome upon measles virus (MV) infection. The novelty about this study is the side-by side comparison of different strains from the same virus, which has not been done at the proteome level for any other virus including MV. We used different virus strains including a vaccine strain, wild type isolates derived from MV-infected patients as well as a Vero cell adapted strain, which serves as an intermediate between vaccine and wild type strain. We observed differences between vaccine and wild type strains as well as common features between different wild type strains. Perhaps one of the most surprising findings was that differences did not only occur between wild type and vaccine or Vero cell adapted strains but also between different wild type strains. In fact our study suggests that besides the cytokeratin and the IFN system wild type viruses seem to differ as much among each other than from vaccine strains. Thus our results are suggestive of complex and diverse virus-host interactions which differ considerably between different wild type strains. Our data indicate that interstrain differences are prominent and have so far been neglected by proteomics studies.