31 results on '"Lucas M Ferreri"'
Search Results
2. Timing of exposure is critical in a highly sensitive model of SARS-CoV-2 transmission.
- Author
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Ketaki Ganti, Lucas M Ferreri, Chung-Young Lee, Camden R Bair, Gabrielle K Delima, Kate E Holmes, Mehul S Suthar, and Anice C Lowen
- Subjects
Immunologic diseases. Allergy ,RC581-607 ,Biology (General) ,QH301-705.5 - Abstract
Transmission efficiency is a critical factor determining the size of an outbreak of infectious disease. Indeed, the propensity of SARS-CoV-2 to transmit among humans precipitated and continues to sustain the COVID-19 pandemic. Nevertheless, the number of new cases among contacts is highly variable and underlying reasons for wide-ranging transmission outcomes remain unclear. Here, we evaluated viral spread in golden Syrian hamsters to define the impact of temporal and environmental conditions on the efficiency of SARS-CoV-2 transmission through the air. Our data show that exposure periods as brief as one hour are sufficient to support robust transmission. However, the timing after infection is critical for transmission success, with the highest frequency of transmission to contacts occurring at times of peak viral load in the donor animals. Relative humidity and temperature had no detectable impact on transmission when exposures were carried out with optimal timing and high inoculation dose. However, contrary to expectation, trends observed with sub-optimal exposure timing and lower inoculation dose suggest improved transmission at high relative humidity or high temperature. In sum, among the conditions tested, our data reveal the timing of exposure to be the strongest determinant of SARS-CoV-2 transmission success and implicate viral load as an important driver of transmission.
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- 2022
- Full Text
- View/download PDF
3. Characterizing Emerging Canine H3 Influenza Viruses.
- Author
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Luis Martinez-Sobrido, Pilar Blanco-Lobo, Laura Rodriguez, Theresa Fitzgerald, Hanyuan Zhang, Phuong Nguyen, Christopher S Anderson, Jeanne Holden-Wiltse, Sanjukta Bandyopadhyay, Aitor Nogales, Marta L DeDiego, Brian R Wasik, Benjamin L Miller, Carole Henry, Patrick C Wilson, Mark Y Sangster, John J Treanor, David J Topham, Lauren Byrd-Leotis, David A Steinhauer, Richard D Cummings, Jasmina M Luczo, Stephen M Tompkins, Kaori Sakamoto, Cheryl A Jones, John Steel, Anice C Lowen, Shamika Danzy, Hui Tao, Ashley L Fink, Sabra L Klein, Nicholas Wohlgemuth, Katherine J Fenstermacher, Farah El Najjar, Andrew Pekosz, Lauren Sauer, Mitra K Lewis, Kathryn Shaw-Saliba, Richard E Rothman, Zhen-Ying Liu, Kuan-Fu Chen, Colin R Parrish, Ian E H Voorhees, Yoshihiro Kawaoka, Gabriele Neumann, Shiho Chiba, Shufang Fan, Masato Hatta, Huihui Kong, Gongxun Zhong, Guojun Wang, Melissa B Uccellini, Adolfo García-Sastre, Daniel R Perez, Lucas M Ferreri, Sander Herfst, Mathilde Richard, Ron Fouchier, David Burke, David Pattinson, Derek J Smith, Victoria Meliopoulos, Pamela Freiden, Brandi Livingston, Bridgett Sharp, Sean Cherry, Juan Carlos Dib, Guohua Yang, Charles J Russell, Subrata Barman, Richard J Webby, Scott Krauss, Angela Danner, Karlie Woodard, Malik Peiris, R A P M Perera, M C W Chan, Elena A Govorkova, Bindumadhav M Marathe, Philippe N Q Pascua, Gavin Smith, Yao-Tsun Li, Paul G Thomas, and Stacey Schultz-Cherry
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Immunologic diseases. Allergy ,RC581-607 ,Biology (General) ,QH301-705.5 - Abstract
The continual emergence of novel influenza A strains from non-human hosts requires constant vigilance and the need for ongoing research to identify strains that may pose a human public health risk. Since 1999, canine H3 influenza A viruses (CIVs) have caused many thousands or millions of respiratory infections in dogs in the United States. While no human infections with CIVs have been reported to date, these viruses could pose a zoonotic risk. In these studies, the National Institutes of Allergy and Infectious Diseases (NIAID) Centers of Excellence for Influenza Research and Surveillance (CEIRS) network collaboratively demonstrated that CIVs replicated in some primary human cells and transmitted effectively in mammalian models. While people born after 1970 had little or no pre-existing humoral immunity against CIVs, the viruses were sensitive to existing antivirals and we identified a panel of H3 cross-reactive human monoclonal antibodies (hmAbs) that could have prophylactic and/or therapeutic value. Our data predict these CIVs posed a low risk to humans. Importantly, we showed that the CEIRS network could work together to provide basic research information important for characterizing emerging influenza viruses, although there were valuable lessons learned.
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- 2020
- Full Text
- View/download PDF
4. Swine influenza A virus isolates containing the pandemic H1N1 origin matrix gene elicit greater disease in the murine model
- Author
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Shelly J. Curran, Emily F. Griffin, Lucas M. Ferreri, Constantinos S. Kyriakis, Elizabeth W. Howerth, Daniel R. Perez, and S. Mark Tompkins
- Subjects
influenza virus ,pathogenesis ,swine influenza ,mouse model ,matrix gene ,pandemic influenza ,Microbiology ,QR1-502 - Abstract
ABSTRACTSince the 1990s, endemic North American swine influenza A viruses (swFLUAVs) contained an internal gene segment constellation, the triple reassortment internal gene (TRIG) cassette. In 2009, the H1N1 pandemic (pdmH1N1) virus spilled back into swine but did not become endemic. However, the pdmH1N1 contributed the matrix gene (pdmM) to the swFLUAVs circulating in the pig population, which replaced the classical swine matrix gene (swM) found in the TRIG cassette, suggesting the pdmM has a fitness benefit. Others have shown that swFLUAVs containing the pdmM have greater transmission efficiency compared to viruses containing the swM gene segment. We hypothesized that the matrix (M) gene could also affect disease and utilized two infection models, resistant BALB/c and susceptible DBA/2 mice, to assess pathogenicity. We infected BALB/c and DBA/2 mice with H1 and H3 swFLUAVs containing the swM or pdmM and measured lung virus titers, morbidity, mortality, and lung histopathology. H1 influenza strains containing the pdmM gene caused greater morbidity and mortality in resistant and susceptible murine strains, while H3 swFLUAVs caused no clinical disease. However, both H1 and H3 swFLUAVs containing the pdmM replicated to higher viral titers in the lungs and pdmM containing H1 viruses induced greater histological changes compared to swM H1 viruses. While the surface glycoproteins and other gene segments may contribute to swFLUAV pathogenicity in mice, these data suggest that the origin of the matrix gene also contributes to pathogenicity of swFLUAV in mice, although we must be cautious in translating these conclusions to their natural host, swine.IMPORTANCEThe 2009 pandemic H1N1 virus rapidly spilled back into North American swine, reassorting with the already genetically diverse swFLUAVs. Notably, the M gene segment quickly replaced the classical M gene segment, suggesting a fitness benefit. Here, using two murine models of infection, we demonstrate that swFLUAV isolates containing the pandemic H1N1 origin M gene caused increased disease compared to isolates containing the classical swine M gene. These results suggest that, in addition to other influenza virus gene segments, the swFLUAV M gene segment contributes to pathogenesis in mammals.
- Published
- 2024
- Full Text
- View/download PDF
5. South American H4N2 influenza A virus improved replication in chicken trachea after low number of passages
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Lucas M. Ferreri, Silvia Carnaccini, Valeria Olivera, Ariel Pereda, Daniela Rajao, and Daniel R. Perez
- Subjects
General Veterinary - Abstract
Introduction of influenza A viruses (FLUAV) into poultry from waterfowl is frequent, producing economic burden and increasing the probability of human infections. We have previously described the presence of FLUAV in wild birds in Argentina with unique evolutionary trajectories belonging to a South American lineage different from the North American and Eurasian lineages. Adaptability of this South American lineage FLUAV to poultry species is still poorly understood. In the present report, we evaluated the capacity of an H4N2 FLUAV from the South American lineage to adapt to chickens after low number of passages. We found that five mutations were acquired after five passages in 3-days-old chickens. These mutations produced a virus with better infectivity in ex vivo trachea explants but overall lower infection in lung explants. Infection of 3-week-old chickens persisted for a longer period and was detected in more tissues than the parental virus, suggesting adaptation of the H4N2 influenza A virus to chicken.
- Published
- 2023
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6. Antigenic mapping of the hemagglutinin of the H9 subtype influenza A viruses using sera from Japanese quail (Coturnix c. japonica)
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Silvia Carnaccini, C. Joaquín Cáceres, L. Claire Gay, Lucas M. Ferreri, Eugene Skepner, David F. Burke, Ian H. Brown, Ginger Geiger, Adebimpe Obadan, Daniela S. Rajao, Nicola S. Lewis, and Daniel R. Perez
- Abstract
Influenza A viruses (FLUAV) of the H9N2 subtype are zoonotic pathogens that cause significant economic damage to the poultry industry. Vaccination to prevent and control H9N2 infections in poultry is widely employed in the Middle East and Asia. We used phylogenetics and antigenic analysis to study the antigenic properties of the H9 hemagglutinin (HA) using sera produced in Japanese quail (Coturnix c. japonica). Consensus HA1 sequences were generated to capture antigenic diversity among isolates. We constructed chimeric H9N2 viruses containing the HA1 of each consensus sequence on a constant isogenic backbone. The resulting viruses were used to generate antisera from quail, a common and significant minor poultry species whose anti-HA response profiles remain poorly defined. Antigenic maps were generated by plotting the cross-hemagglutination inhibition (HI) data from the panel of quail sera against the chimeric constructs and 51 H9 field isolates. The chimeric antigens were divided into four different antigenic profiles (cyan, blue, orange, and red). Site-directed mutagenesis analysis showed 9 amino acid positions of antigenic relevance. Substitutions at amino acid positions 149, 150, and 180 (H9 HA numbering) had relatively significant impact on HI activity using quail sera. Substitutions E180A and R131K/E180A led to the most significant antigenic change transitions. This study provides insights into the antigenic profile of H9 FLUAVs, with important implications for understanding antigenic drift and improving vaccine development for use in minor poultry species.IMPORTANCEDetermining the relevant amino acids involved in antigenic drift on the surface protein hemagglutinin (HA) is critical to understand influenza virus evolution and efficient assessment of vaccine strains relative to current circulating strains. We used antigenic cartography to generate an antigenic map of the H9 HA using sera produced in one of the most relevant minor poultry species, Japanese quail. Key antigenic positions were identified and tested to confirm their impact on the antigenic profile. This work provides a better understanding of the antigenic diversity of the H9 HA as it relates to reactivity to quail sera and will facilitate a rational approach for selecting more efficacious vaccines against poultry-origin H9 influenza viruses in minor poultry species.
- Published
- 2023
7. Mutation E48K in PB1 Polymerase Subunit Improves Stability of a Candidate Live Attenuated Influenza B Virus Vaccine
- Author
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Jongsuk Mo, Stivalis Cardenas-Garcia, Jefferson J. S. Santos, Lucas M. Ferreri, C. Joaquín Cáceres, Ginger Geiger, Daniel R. Perez, and Daniela S. Rajao
- Subjects
Influenza B ,LAIV ,PB1 ,vaccine stability ,Medicine - Abstract
Influenza B virus (IBV) is a major respiratory pathogen of humans, particularly in the elderly and children, and vaccines are the most effective way to control it. In previous work, incorporation of two mutations (E580G, S660A) along with the addition of an HA epitope tag in the PB1 segment of B/Brisbane/60/2008 (B/Bris) resulted in an attenuated strain that was safe and effective as a live attenuated vaccine. A third attempted mutation (K391E) in PB1 was not always stable. Interestingly, viruses that maintained the K391E mutation were associated with the mutation E48K. To explore the contribution of the E48K mutation to stability of the K391E mutation, a vaccine candidate was generated by inserting both mutations, along with attenuating mutations E580G and S660A, in PB1 of B/Bris (B/Bris PB1att 4M). Serial passages of the B/Bris PB1att 4M vaccine candidate in eggs and MDCK indicated high stability. In silico structural analysis revealed a potential interaction between amino acids at positions 48 and 391. In mice, B/Bris PB1att 4M was safe and provided complete protection against homologous challenge. These results confirm the compensatory effect of mutation E48K to stabilize the K391E mutation, resulting in a safer, yet still protective, IBV LAIV vaccine.
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- 2021
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8. Development of a Novel Live Attenuated Influenza A Virus Vaccine Encoding the IgA-Inducing Protein
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C. Joaquín Cáceres, Stivalis Cardenas-Garcia, Aarti Jain, L. Claire Gay, Silvia Carnaccini, Brittany Seibert, Lucas M. Ferreri, Ginger Geiger, Algimantas Jasinskas, Rie Nakajima, Daniela S. Rajao, Irina Isakova-Sivak, Larisa Rudenko, Amy L. Vincent, D. Huw Davies, and Daniel R. Perez
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LAIV ,influenza ,HA ,IGIP ,IgA ,IgG ,Medicine - Abstract
Live attenuated influenza virus (LAIV) vaccines elicit a combination of systemic and mucosal immunity by mimicking a natural infection. To further enhance protective mucosal responses, we incorporated the gene encoding the IgA-inducing protein (IGIP) into the LAIV genomes of the cold-adapted A/Leningrad/134/17/57 (H2N2) strain (caLen) and the experimental attenuated backbone A/turkey/Ohio/313053/04 (H3N2) (OH/04att). Incorporation of IGIP into the caLen background led to a virus that grew poorly in prototypical substrates. In contrast, IGIP in the OH/04att background (IGIP-H1att) virus grew to titers comparable to the isogenic backbone H1att (H1N1) without IGIP. IGIP-H1att- and H1caLen-vaccinated mice were protected against lethal challenge with a homologous virus. The IGIP-H1att vaccine generated robust serum HAI responses in naïve mice against the homologous virus, equal or better than those obtained with the H1caLen vaccine. Analyses of IgG and IgA responses using a protein microarray revealed qualitative differences in humoral and mucosal responses between vaccine groups. Overall, serum and bronchoalveolar lavage samples from the IGIP-H1att group showed trends towards increased stimulation of IgG and IgA responses compared to H1caLen samples. In summary, the introduction of genes encoding immunomodulatory functions into a candidate LAIV can serve as natural adjuvants to improve overall vaccine safety and efficacy.
- Published
- 2021
- Full Text
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9. Epistasis reduces fitness costs of influenza A virus escape from stem-binding antibodies
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Chung-Young Lee, Vedhika Raghunathan, C. Joaquin Caceres, Ginger Geiger, Brittany Seibert, Flavio Cargnin Faccin, L. Claire Gay, Lucas M. Ferreri, Drishti Kaul, Jens Wrammert, Gene S. Tan, Daniel R. Perez, and Anice C. Lowen
- Subjects
Multidisciplinary - Abstract
The hemagglutinin (HA) stem region is a major target of universal influenza vaccine efforts owing to the presence of highly conserved epitopes across multiple influenza A virus (IAV) strains and subtypes. To explore the potential impact of vaccine-induced immunity targeting the HA stem, we examined the fitness effects of viral escape from stem-binding broadly neutralizing antibodies (stem-bnAbs). Recombinant viruses containing each individual antibody escape substitution showed diminished replication compared to wild-type virus, indicating that stem-bnAb escape incurred fitness costs. A second-site mutation in the HA head domain (N129D; H1 numbering) reduced the fitness effects observed in primary cell cultures and likely enabled the selection of escape mutations. Functionally, this putative permissive mutation increased HA avidity for its receptor. These results suggest a mechanism of epistasis in IAV, wherein modulating the efficiency of attachment eases evolutionary constraints imposed by the requirement for membrane fusion. Taken together, the data indicate that viral escape from stem-bnAbs is costly but highlights the potential for epistatic interactions to enable evolution within the functionally constrained HA stem domain.
- Published
- 2023
10. Live attenuated influenza A virus vaccine expressing an IgA-inducing protein protects pigs against replication and transmission
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Daniela S. Rajao, Giovana C. Zanella, Meghan Wymore Brand, Shehroz Khan, Michael E. Miller, Lucas M. Ferreri, C. Joaquin Caceres, Stivalis Cadernas-Garcia, Carine K. Souza, Tavis K. Anderson, Phillip C. Gauger, Amy L. Vincent Baker, and Daniel R. Perez
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General Medicine - Abstract
IntroductionThe rapid evolution of influenza A viruses (FLUAV) complicates disease control for animal and public health. Although vaccination is an effective way to control influenza, available vaccines for use in swine result in limited protection against the antigenically distinct FLUAV that currently co-circulate in pigs. Vaccines administered parenterally usually stimulate IgG antibodies but not strong mucosal IgA or cell-mediated responses, which are typically more cross-reactive.MethodsWe developed a live attenuated influenza virus (LAIV) vaccine containing IgA-inducing protein (IGIP) as a molecular marker and immunomodulator. This Flu-IGIP vaccine was tested in a bivalent formulation (H1N1 and H3N2) against challenge with antigenically drifted viruses in pigs. Pigs were vaccinated intranasally with either a bivalent Flu-IGIP or a bivalent Flu-att (control without IGIP) and boosted two weeks later. Three weeks post boost, pigs were challenged with antigenically drifted H1N1 or H3N2 virus.ResultsVaccinated pigs had increased numbers of influenza-specific IgA-secreting cells in PBMC two weeks post boost and higher numbers of total and influenza-specific IgA-secreting cells in bronchoalveolar lavage fluid (BALF) 5 days post inoculation (dpi) compared to naïve pigs. Pigs vaccinated with both Flu-IGIP and Flu-att shed significantly less virus after H1N1 or H3N2 challenge compared to non-vaccinated pigs. Vaccination with Flu-att reduced respiratory transmission, while Flu-IGIP fully blocked transmission regardless of challenge virus. Both Flu-IGIP and Flu-att vaccines reduced virus replication in the lungs and lung lesions after inoculation with either virus. IgG and IgA levels in BALF and nasal wash of vaccinated pigs were boosted after inoculation as soon as 5 dpi and remained high at 14 dpi.ConclusionOur results indicate that Flu-IGIP leads to protection from clinical signs, replication and shedding after antigenically drifted influenza virus infection.
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- 2023
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11. Transmission of Human Influenza A Virus in Pigs Selects for Adaptive Mutations on the HA Gene
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Jong-suk Mo, Eugenio J. Abente, Matias Cardenas Perez, Troy C. Sutton, Brianna Cowan, Lucas M. Ferreri, Ginger Geiger, Phillip C. Gauger, Daniel R. Perez, Amy L. Vincent Baker, and Daniela S. Rajao
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Swine Diseases ,Swine ,Immunology ,Hemagglutinin Glycoproteins, Influenza Virus ,Microbiology ,Orthomyxoviridae Infections ,Genetic Diversity and Evolution ,Influenza A virus ,Virology ,Insect Science ,Influenza, Human ,Mutation ,Animals ,Humans ,Reassortant Viruses - Abstract
Influenza A viruses (FLUAV) cause respiratory diseases in many host species, including humans and pigs. The spillover of FLUAV between swine and humans has been a concern for both public health and the swine industry. With the emergence of the triple reassortant internal gene (TRIG) constellation, establishment of human-origin FLUAVs in pigs has become more common, leading to increased viral diversity. However, little is known about the adaptation processes that are needed for a human-origin FLUAV to transmit and become established in pigs. We generated a reassortant FLUAV (VIC11pTRIG) containing surface gene segments from a human FLUAV strain and internal gene segments from the 2009 pandemic and TRIG FLUAV lineages and demonstrated that it can replicate and transmit in pigs. Sequencing and variant analysis identified three mutants that emerged during replication in pigs, which were mapped near the receptor binding site of the hemagglutinin (HA). The variants replicated more efficiently in differentiated swine tracheal cells compared to the virus containing the wildtype human-origin HA, and one of them was present in all contact pigs. These results show that variants are selected quickly after replication of human-origin HA in pigs, leading to improved fitness in the swine host, likely contributing to transmission. IMPORTANCE Influenza A viruses cause respiratory disease in several species, including humans and pigs. The bidirectional transmission of FLUAV between humans and pigs plays a significant role in the generation of novel viral strains, greatly impacting viral epidemiology. However, little is known about the evolutionary processes that allow human FLUAV to become established in pigs. In this study, we generated reassortant viruses containing human seasonal HA and neuraminidase (NA) on different constellations of internal genes and tested their ability to replicate and transmit in pigs. We demonstrated that a virus containing a common internal gene constellation currently found in U.S. swine was able to transmit efficiently via the respiratory route. We identified a specific amino acid substitution that was fixed in the respiratory contact pigs that was associated with improved replication in primary swine tracheal epithelial cells, suggesting it was crucial for the transmissibility of the human virus in pigs.
- Published
- 2022
12. Epistasis reduces fitness costs of influenza A virus escape from stem-binding antibodies
- Author
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Chung-Young Lee, C. Joaquin Caceres, Ginger Geiger, Brittany Seibert, Flavio Cargnin Faccin, L. Claire Gay, Lucas M. Ferreri, Drishti Kaul, Jens Wrammert, Gene S. Tan, Daniel R. Perez, and Anice C. Lowen
- Abstract
The hemagglutinin (HA) stem region is a major target of universal influenza vaccine efforts owing to the presence of highly conserved epitopes across multiple influenza A virus strains and subtypes. To explore the potential impact of vaccine-induced immunity targeting the HA stem, we examined the fitness effects of viral escape from stem-binding broadly neutralizing antibodies (stem-bnAbs). Recombinant viruses containing each individual antibody escape substitution showed diminished replication compared to wild-type virus, indicating that stem-bnAb escape incurred fitness costs. A second-site mutation in the HA head domain (N133D) reduced the fitness effects observed in primary cell cultures and likely enabled the selection of escape mutations. This putative permissive mutation was not, however, sufficient to ease fitness costs in a ferret transmission model. Taken together, these data suggest that viral escape from stem-bnAbs is costly but highlight the potential for epistatic interactions to enable evolution within the functionally constrained HA stem domain.
- Published
- 2022
13. Influenza A virus reassortment in mammals gives rise to genetically distinct within-host sub-populations
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Ketaki Ganti, Anish Bagga, Silvia Carnaccini, Lucas M. Ferreri, Ginger Geiger, C. Joaquin Caceres, Brittany Seibert, Yonghai Li, Liping Wang, Taeyong Kwon, Yuhao Li, Igor Morozov, Wenjun Ma, Juergen A. Richt, Daniel R. Perez, Katia Koelle, and Anice C. Lowen
- Subjects
Mammals ,Swine Diseases ,Multidisciplinary ,Swine ,Guinea Pigs ,Ferrets ,General Physics and Astronomy ,General Chemistry ,General Biochemistry, Genetics and Molecular Biology ,Influenza A Virus, H1N1 Subtype ,Orthomyxoviridae Infections ,Influenza A virus ,Influenza, Human ,Animals ,Humans ,Reassortant Viruses - Abstract
Influenza A virus (IAV) genetic exchange through reassortment has the potential to accelerate viral evolution and has played a critical role in the generation of multiple pandemic strains. For reassortment to occur, distinct viruses must co-infect the same cell. The spatio-temporal dynamics of viral dissemination within an infected host therefore define opportunity for reassortment. Here, we used wild type and synonymously barcoded variant viruses of a pandemic H1N1 strain to examine the within-host viral dynamics that govern reassortment in guinea pigs, ferrets and swine. The first two species are well-established models of human influenza, while swine are a natural host and a frequent conduit for cross-species transmission and reassortment. Our results show reassortment to be pervasive in all three hosts but less frequent in swine than in ferrets and guinea pigs. In ferrets, tissue-specific differences in the opportunity for reassortment are also evident, with more reassortants detected in the nasal tract than the lower respiratory tract. While temporal trends in viral diversity are limited, spatial patterns are clear, with heterogeneity in the viral genotypes detected at distinct anatomical sites revealing extensive compartmentalization of reassortment and replication. Our data indicate that the dynamics of viral replication in mammals allow diversification through reassortment but that the spatial compartmentalization of variants likely shapes their evolution and onward transmission.
- Published
- 2022
14. Collective interactions augment influenza A virus replication in a host-dependent manner
- Author
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Gene S. Tan, Silvia Carnaccini, Maria C. White, Anice C. Lowen, Lucas M. Ferreri, Ketaki Ganti, Brett E. Pickett, Kara L. Phipps, Miglena Manandhar, Daniel R. Perez, Chung-Young Lee, and Nathan T. Jacobs
- Subjects
Microbiology (medical) ,viruses ,Immunology ,Cell ,Gene Expression ,Biology ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Microbiology ,Genome ,Antiviral Agents ,Virus ,Article ,03 medical and health sciences ,Gene mapping ,Transcription (biology) ,Genetics ,medicine ,Influenza A virus ,Gene ,030304 developmental biology ,0303 health sciences ,030306 microbiology ,Cell Biology ,medicine.disease ,Phenotype ,Virology ,Complementation ,medicine.anatomical_structure ,Viral replication ,Coinfection - Abstract
Infection with a single influenza A virus (IAV) is only rarely sufficient to initiate productive infection. Here, we exploit both single-cell approaches and whole-animal systems to show that IAV reliance on multiple infection can form an important species barrier to infection. Namely, we find that H9N2 subtype viruses representative of those circulating widely at the poultry-human interface exhibit acute dependence on collective interactions in mammalian systems. This need for multiple infection is greatly reduced in the natural host. Quantification of incomplete viral genomes showed that their complementation accounts for the more moderate reliance on coinfection seen in avian cells, but not the added reliance seen in mammalian cells. This finding suggests an additional form of virus-virus interaction is needed to support infection in mammalian cells. Genetic mapping implicated the PA gene segment as a major driver of this phenotype and quantification of viral RNA synthesis indicated that both replication and transcription were affected. These findings indicate that multiple distinct mechanisms underlie IAV reliance on multiple infection and underscore the importance of virus-virus interactions in IAV infection, evolution and emergence.
- Published
- 2020
15. Timing of exposure is critical in a highly sensitive model of SARS-CoV-2 transmission
- Author
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Ketaki Ganti, Lucas M. Ferreri, Chung-Young Lee, Camden R. Bair, Gabrielle K. Delima, Kate E. Holmes, Mehul S. Suthar, and Anice C. Lowen
- Subjects
Mesocricetus ,SARS-CoV-2 ,Immunology ,COVID-19 ,Viral Load ,Microbiology ,Virology ,Cricetinae ,Genetics ,Animals ,Humans ,Parasitology ,Molecular Biology ,Pandemics - Abstract
Transmission efficiency is a critical factor determining the size of an outbreak of infectious disease. Indeed, the propensity of SARS-CoV-2 to transmit among humans precipitated and continues to sustain the COVID-19 pandemic. Nevertheless, the number of new cases among contacts is highly variable and underlying reasons for wide-ranging transmission outcomes remain unclear. Here, we evaluated viral spread in golden Syrian hamsters to define the impact of temporal and environmental conditions on the efficiency of SARS-CoV-2 transmission through the air. Our data show that exposure periods as brief as one hour are sufficient to support robust transmission. However, the timing after infection is critical for transmission success, with the highest frequency of transmission to contacts occurring at times of peak viral load in the donor animals. Relative humidity and temperature had no detectable impact on transmission when exposures were carried out with optimal timing and high inoculation dose. However, contrary to expectation, trends observed with sub-optimal exposure timing and lower inoculation dose suggest improved transmission at high relative humidity or high temperature. In sum, among the conditions tested, our data reveal the timing of exposure to be the strongest determinant of SARS-CoV-2 transmission success and implicate viral load as an important driver of transmission.
- Published
- 2021
16. Intra- and inter-host evolution of H9N2 influenza A virus in Japanese quail
- Author
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Lucas M Ferreri, Ginger Geiger, Brittany Seibert, Adebimpe Obadan, Daniela Rajao, Anice C Lowen, and Daniel R Perez
- Subjects
Virology ,Microbiology - Abstract
Influenza A viruses (IAVs) are constantly evolving. Crucial steps in the infection cycle, such as sialic acid (SA) receptor binding on the host cell surface, can either promote or hamper the emergence of new variants. We previously assessed the relative fitness in Japanese quail of H9N2 variant viruses differing at a single amino acid position, residue 216 in the hemagglutinin (HA) viral surface protein. This site is known to modulate SA recognition. Our prior study generated a valuable set of longitudinal samples from quail transmission groups where the inoculum comprised different mixed populations of HA 216 variant viruses. Here, we leveraged these samples to examine the evolutionary dynamics of viral populations within and between inoculated and naïve contact quails. We found that positive selection dominated HA gene evolution, but fixation of the fittest variant depended on the competition mixture. Analysis of the whole genome revealed further evidence of positive selection acting both within and between hosts. Positive selection drove fixation of variants in non-HA segments within inoculated and contact quails. Importantly, transmission bottlenecks were modulated by the molecular signature at HA 216, revealing viral receptor usage as a determinant of transmitted diversity. Overall, we show that selection strongly shaped the evolutionary dynamics within and between quails. These findings support the notion that selective processes act effectively on IAV populations in poultry hosts, facilitating rapid viral evolution in this ecological niche.
- Published
- 2021
17. Mutation E48K in PB1 Polymerase Subunit Improves Stability of a Candidate Live Attenuated Influenza B Virus Vaccine
- Author
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Lucas M. Ferreri, C. Joaquín Cáceres, Jefferson Santos, Daniela S. Rajao, Ginger Geiger, Stivalis Cardenas-Garcia, Daniel R. Perez, and Jong-Suk Mo
- Subjects
0301 basic medicine ,Protein subunit ,Immunology ,vaccine stability ,Biology ,Influenza B ,Virus ,Epitope ,Article ,03 medical and health sciences ,0302 clinical medicine ,Drug Discovery ,Homologous chromosome ,biochemistry ,Live attenuated influenza vaccine ,Pharmacology (medical) ,030212 general & internal medicine ,Polymerase ,LAIV ,Pharmacology ,Attenuated vaccine ,Virology ,030104 developmental biology ,Infectious Diseases ,PB1 ,Mutation (genetic algorithm) ,biology.protein ,Medicine - Abstract
Influenza B virus (IBV) is a major respiratory pathogen of humans, particularly in the elderly and children, and vaccines are the most effective way to control it. In previous work, incorporation of two mutations (E580G, S660A) along with the addition of an HA epitope tag in the PB1 segment of B/Brisbane/60/2008 (B/Bris) resulted in an attenuated strain that was safe and effective as a live attenuated vaccine. A third attempted mutation (K391E) in PB1 was not always stable. Interestingly, viruses that maintained the K391E mutation were associated with the mutation E48K. To explore the contribution of the E48K mutation to stability of the K391E mutation, a vaccine candidate was generated by inserting both mutations, along with attenuating mutations E580G and S660A, in PB1 of B/Bris (B/Bris PB1att 4M). Serial passages of the B/Bris PB1att 4M vaccine candidate in eggs and MDCK indicated high stability. In silico structural analysis revealed a potential interaction between amino acids at positions 48 and 391. In mice, B/Bris PB1att 4M was safe and provided complete protection against homologous challenge. These results confirm the compensatory effect of mutation E48K to stabilize the K391E mutation, resulting in a safer, yet still protective, IBV LAIV vaccine.
- Published
- 2021
18. Evidence of a fixed internal gene constellation in influenza A viruses isolated from wild birds in Argentina (2006–2016)
- Author
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Ariel Pereda, Ana S. Gonzalez-Reiche, Agustina Rimondi, Valeria Olivera, Martha I. Nelson, Daniel R. Perez, Ginger Geiger, Marcelo Romano, Harm van Bakel, Gabriel J. Castresana, Lucas M. Ferreri, and Julieta Decarre
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0301 basic medicine ,Genes, Viral ,Epidemiology ,Lineage (evolution) ,Waterfowl ,Reassortment ,lcsh:QR1-502 ,Influenza A Virus, H7N9 Subtype ,medicine.disease_cause ,lcsh:Microbiology ,purl.org/becyt/ford/1 [https] ,Feces ,Influenza A Virus, H1N1 Subtype ,Cloaca ,Drug Discovery ,Influenza A virus ,Phylogeny ,ARGENTINA ,Phylogenetic tree ,biology ,virus diseases ,General Medicine ,Infectious Diseases ,INFLUENZA ,Aves ,CIENCIAS NATURALES Y EXACTAS ,Reassortant Viruses ,Immunology ,Argentina ,Hemagglutinins, Viral ,Neuraminidase ,Zoology ,Hemagglutinin (influenza) ,Animals, Wild ,Genome, Viral ,Real-Time Polymerase Chain Reaction ,ECOLOGY ,Microbiology ,Article ,Virus ,lcsh:Infectious and parasitic diseases ,Ciencias Biológicas ,Birds ,Evolution, Molecular ,03 medical and health sciences ,Avian Influenzavirus ,Virology ,medicine ,Animals ,Virus de la Influenza Aviar ,lcsh:RC109-216 ,purl.org/becyt/ford/1.6 [https] ,Disease Reservoirs ,Aves Acuáticas ,EVOLUTION ,Divergent evolution ,Filogenia ,030104 developmental biology ,Genes ,Influenza in Birds ,biology.protein ,Parasitology ,Virología - Abstract
Wild aquatic birds are the major reservoir of influenza A virus. Cloacal swabs and feces samples (n = 6595) were collected from 62 bird species in Argentina from 2006 to 2016 and screened for influenza A virus. Full genome sequencing of 15 influenza isolates from 6 waterfowl species revealed subtypes combinations that were previously described in South America (H1N1, H4N2, H4N6 (n = 3), H5N3, H6N2 (n = 4), and H10N7 (n = 2)), and new ones not previously identified in the region (H4N8, H7N7 and H7N9). Notably, the internal gene segments of all 15 Argentine isolates belonged to the South American lineage, showing a divergent evolution of these viruses in the Southern Hemisphere. Time-scaled phylogenies indicated that South American gene segments diverged between ~ 30 and~ 140 years ago from the most closely related influenza lineages, which include the avian North American (PB1, HA, NA, MP, and NS-B) and Eurasian lineage (PB2), and the equine H3N8 lineage (PA, NP, and NS-A). Phylogenetic analyses of the hemagglutinin and neuraminidase gene segments of the H4, H6, and N8 subtypes revealed recent introductions and reassortment between viruses from the Northern and Southern Hemispheres in the Americas. Remarkably and despite evidence of recent hemagglutinin and neuraminidase subtype introductions, the phylogenetic composition of internal gene constellation of these influenza A viruses has remained unchanged. Considering the extended time and the number of sampled species of the current study, and the paucity of previously available data, our results contribute to a better understanding of the ecology and evolution of influenza virus in South America. Fil: Rimondi, Agustina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Virología; Argentina Fil: Gonzalez Reiche, Ana S.. University of Georgia; Estados Unidos Fil: Olivera, Valeria. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Virología; Argentina Fil: Decarre, Julieta. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Recursos Naturales. Instituto de Recursos Biológicos; Argentina Fil: Castresana, Gabriel J.. Gobierno de la Provincia de Buenos Aires. Organismo Provincial Para El Desarrollo Sostenible.; Argentina Fil: Romano, Marcelo. Centro de Investigaciones en Biodiversidad y Ambiente; Argentina Fil: Nelson, Martha I.. National Institutes of Health; Estados Unidos Fil: van Bakel, Harm. Icahn School of Medicine at Mount Sina; Estados Unidos Fil: Pereda, Ariel Julián. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Patobiología; Argentina Fil: Ferreri, Lucas. University of Georgia; Estados Unidos. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Geiger, Ginger. University of Georgia; Estados Unidos Fil: Perez, Daniel R.. University of Georgia; Estados Unidos
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- 2018
19. Silent infection of B and CD8 + T lymphocytes by Influenza A virus in children with tonsillar hypertrophy
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Eurico Arruda, Marjorie Cornejo Pontelli, Fabiana Cardoso Pereira Valera, Daniel Macedo de Melo Jorge, Miria Ferreira Criado, Italo A. Castro, Wilma Terezinha Anselmo-Lima, Ricardo S. Cardoso, Edwin Tamashiro, Lucas R. Carenzi, Bruna Lais Santos de Jesus, Daniel R. Perez, Lucas M. Ferreri, and Ronaldo B. Martins
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0303 health sciences ,education.field_of_study ,030306 microbiology ,Immunology ,Population ,Biology ,VIROLOGIA ,medicine.disease_cause ,Microbiology ,Virology ,Virus ,03 medical and health sciences ,Immune system ,Lymphatic system ,Viral replication ,Insect Science ,Influenza A virus ,medicine ,Tissue tropism ,Viral shedding ,education ,030304 developmental biology - Abstract
Influenza A viruses (IAVs) cause more than 2 million annual episodes of seasonal acute respiratory infections (ARI) and approximately 500,000 deaths worldwide. Depending on virus strain and host immune status, acute infections by IAV may reach sites other than the respiratory tract. In the present study, IAV RNA and antigens were searched for in tissues of palatine tonsils and adenoids removed from patients without ARI symptoms. A real-time reverse transcriptase PCR (RT-PCR) screening revealed that 8 tissue samples from 7 patients out of 103 were positive for IAV. Positive samples were subjected to next-generation sequencing (NGS) and 3 of 8 tissues yielded complete IAV pH1N1 genomes, whereas in 5 samples, the PB1 gene was not fully assembled. Phylogenetic analysis placed tonsil-derived IAV in clusters clearly segregated from contemporaneous Brazilian viruses. Flow cytometry of dispersed tissue fragments and serial immunohistochemistry of paraffin-embedded sections of naturally infected biopsies indicated that CD20+ B lymphocytes, CD8+ T lymphocytes, and CD11c+ cells are susceptible to IAV infection. We sought to investigate whether these lymphoid tissues could be sites of viral replication and sources of viable virus particles. MDCK cells were inoculated with tissue lysates, enabling recovery of one IAV isolate confirmed by immunofluorescence, reverse transcriptase quantitative PCR (RT-qPCR), and NGS. The data indicate that lymphoid tissues not only harbor expression of IAV proteins but also contain infectious virus. Asymptomatic long-term infection raises the possibility of IAV shedding from tonsils, which may have an impact on host-to-host transmission.IMPORTANCE Influenza A virus (IAV) infections are important threats to human health worldwide. Although extensively studied, some aspects of virus pathogenesis and tissue tropism remain unclear. Here, by different strategies, we describe the asymptomatic infection of human lymphoid organs by IAV in children. Our results indicate that IAV was not only detected and isolated from human tonsils but displayed unique genetic features in comparison with those of contemporaneous IAVs circulating in Brazil and detected in swabs and nasal washes. Inside the tissue microenvironment, immune cells were shown to be carrying IAV antigens, especially B and T CD8+ lymphocytes. Taken together, these results suggest that human lymphoid tissues can be sites of silent IAV infections with possible impact on virus shedding to the population.
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- 2020
20. Characterizing Emerging Canine H3 Influenza Viruses
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Huihui Kong, Laura Rodriguez, Brian R. Wasik, Hanyuan Zhang, Guojun Wang, Yoshihiro Kawaoka, Kuan-Fu Chen, Pamela Freiden, Nicholas Wohlgemuth, David F. Burke, Ashley L. Fink, Sabra L. Klein, Melissa B. Uccellini, Patrick C. Wilson, Karlie Woodard, Sander Herfst, David J. Topham, Angela Danner, Zhen-Ying Liu, Daniel R. Perez, Cheryl A. Jones, John Steel, Philippe Noriel Q. Pascua, Christopher S. Anderson, Anice C. Lowen, Katherine J. Fenstermacher, Lauren Sauer, Victoria A. Meliopoulos, Jeanne Holden-Wiltse, Richard D. Cummings, Yao-Tsun Li, Gabriele Neumann, Malik Peiris, Elena A. Govorkova, Kaori Sakamoto, Michael C. W. Chan, Derek J. Smith, Benjamin L. Miller, Sanjukta Bandyopadhyay, Colin R. Parrish, Subrata Barman, John J. Treanor, Lucas M. Ferreri, Shamika Danzy, Hui Tao, Ian E. H. Voorhees, Ranawaka A.P.M. Perera, Paul G. Thomas, Scott Krauss, David A. Steinhauer, Adolfo García-Sastre, Pilar Blanco-Lobo, Gavin J. D. Smith, Stacey Schultz-Cherry, Phuong T. M. Nguyen, Luis Martinez-Sobrido, Ron A. M. Fouchier, Masato Hatta, Sean Cherry, Brandi Livingston, Marta L. DeDiego, Gongxun Zhong, Mathilde Richard, David J. Pattinson, Mitra Lewis, Bridgett Sharp, Farah El Najjar, Andrew Pekosz, Jasmina M. Luczo, Stephen M. Tompkins, Charles J. Russell, Bindumadhav M. Marathe, Richard E. Rothman, Carole Henry, Lauren Byrd-Leotis, Mark Y. Sangster, Theresa Fitzgerald, Juan Carlos Dib, Shiho Chiba, Shufang Fan, Kathryn Shaw-Saliba, Aitor Nogales, Guohua Yang, Richard J. Webby, Virology, Martinez-Sobrido, Luis [0000-0001-7084-0804], Zhang, Hanyuan [0000-0002-0736-4603], Nguyen, Phuong [0000-0002-8273-730X], Anderson, Christopher S [0000-0002-8560-3438], Holden-Wiltse, Jeanne [0000-0003-2694-7465], Nogales, Aitor [0000-0002-2424-7900], Wasik, Brian R [0000-0001-5442-3883], Miller, Benjamin L [0000-0001-9168-8047], Henry, Carole [0000-0002-5696-527X], Wilson, Patrick C [0000-0002-3537-1245], Topham, David J [0000-0002-9435-8673], Byrd-Leotis, Lauren [0000-0002-7984-0357], Cummings, Richard D [0000-0002-8918-5034], Luczo, Jasmina M [0000-0002-8036-110X], Tompkins, Stephen M [0000-0002-1523-5588], Sakamoto, Kaori [0000-0003-0592-6403], Steel, John [0000-0003-1217-0990], Klein, Sabra L [0000-0002-0730-5224], Wohlgemuth, Nicholas [0000-0002-6450-6452], Fenstermacher, Katherine J [0000-0003-1139-3711], Pekosz, Andrew [0000-0003-3248-1761], Lewis, Mitra K [0000-0002-4737-4961], Chen, Kuan-Fu [0000-0001-7287-9497], Voorhees, Ian E H [0000-0003-3189-1101], García-Sastre, Adolfo [0000-0002-6551-1827], Perez, Daniel R [0000-0002-6569-5689], Ferreri, Lucas M [0000-0002-1069-9500], Herfst, Sander [0000-0001-9866-8903], Richard, Mathilde [0000-0003-0240-9312], Burke, David [0000-0001-8830-3951], Pattinson, David [0000-0003-0001-8203], Smith, Derek J [0000-0002-2393-1890], Freiden, Pamela [0000-0001-6167-180X], Peiris, Malik [0000-0001-8217-5995], Chan, M C W [0000-0001-8174-8405], Govorkova, Elena A [0000-0001-9067-5682], Marathe, Bindumadhav M [0000-0002-9929-7566], Pascua, Philippe N Q [0000-0001-6777-2994], Smith, Gavin [0000-0001-5031-468X], Schultz-Cherry, Stacey [0000-0002-2021-727X], Apollo - University of Cambridge Repository, Voorhees, Ian EH [0000-0003-3189-1101], Chan, MCW [0000-0001-8174-8405], Pascua, Philippe NQ [0000-0001-6777-2994], Anderson, Christopher S. [0000-0002-8560-3438], Wasik, Brian R. [0000-0001-5442-3883], Miller, Benjamin L. [0000-0001-9168-8047], Wilson, Patrick C. [0000-0002-3537-1245], Topham, David J. [0000-0002-9435-8673], Cummings, Richard D. [0000-0002-8918-5034], Luczo, Jasmina M. [0000-0002-8036-110X], Tompkins, Stephen M. [0000-0002-1523-5588], Klein, Sabra L. [0000-0002-0730-5224], Fenstermacher, Katherine J. [0000-0003-1139-3711], Lewis, Mitra K. [0000-0002-4737-4961], Voorhees, Ian E. H. [0000-0003-3189-1101], Perez, Daniel R. [0000-0002-6569-5689], Ferreri, Lucas M. [0000-0002-1069-9500], Smith, Derek J. [0000-0002-2393-1890], Chan, M. C. W. [0000-0001-8174-8405], Govorkova, Elena A. [0000-0001-9067-5682], Marathe, Bindumadhav M. [0000-0002-9929-7566], and Pascua, Philippe N. Q. [0000-0001-6777-2994]
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RNA viruses ,Viral Diseases ,Influenza Viruses ,Pulmonology ,Physiology ,Pathology and Laboratory Medicine ,Communicable Diseases, Emerging ,Biochemistry ,Fluorescence Microscopy ,Basic research ,Zoonoses ,Immune Physiology ,Research article ,Dog Diseases ,Biology (General) ,Enzyme-Linked Immunoassays ,Mammals ,Mice, Inbred BALB C ,Microscopy ,0303 health sciences ,Immune System Proteins ,Viral Vaccine ,030302 biochemistry & molecular biology ,Eukaryota ,Light Microscopy ,Influenza research ,Infectious Diseases ,Influenza A virus ,Mice, Inbred DBA ,Medical Microbiology ,Viral Pathogens ,Viruses ,Vertebrates ,Pathogens ,Research Article ,medicine.medical_specialty ,QH301-705.5 ,Guinea Pigs ,Immunology ,Microbiology ,Antibodies ,Influenza A Virus, H3N8 Subtype ,03 medical and health sciences ,Dogs ,Virology ,Influenza, Human ,Genetics ,medicine ,Animals ,Humans ,Immunoassays ,Microbial Pathogens ,Molecular Biology ,030304 developmental biology ,Medicine and health sciences ,Biology and life sciences ,business.industry ,Influenza A Virus, H3N2 Subtype ,Public health ,Ferrets ,Organisms ,Proteins ,Influenza a ,RC581-607 ,United States ,Influenza ,Viral Replication ,Mice, Inbred C57BL ,Research and analysis methods ,Viral replication ,Respiratory Infections ,Amniotes ,Immunologic Techniques ,Parasitology ,Immunologic diseases. Allergy ,business ,Orthomyxoviruses - Abstract
The continual emergence of novel influenza A strains from non-human hosts requires constant vigilance and the need for ongoing research to identify strains that may pose a human public health risk. Since 1999, canine H3 influenza A viruses (CIVs) have caused many thousands or millions of respiratory infections in dogs in the United States. While no human infections with CIVs have been reported to date, these viruses could pose a zoonotic risk. In these studies, the National Institutes of Allergy and Infectious Diseases (NIAID) Centers of Excellence for Influenza Research and Surveillance (CEIRS) network collaboratively demonstrated that CIVs replicated in some primary human cells and transmitted effectively in mammalian models. While people born after 1970 had little or no pre-existing humoral immunity against CIVs, the viruses were sensitive to existing antivirals and we identified a panel of H3 cross-reactive human monoclonal antibodies (hmAbs) that could have prophylactic and/or therapeutic value. Our data predict these CIVs posed a low risk to humans. Importantly, we showed that the CEIRS network could work together to provide basic research information important for characterizing emerging influenza viruses, although there were valuable lessons learned., Author summary The 2009 influenza pandemic was a stark reminder that ongoing vigilance is critical to protect the public from an influenza pandemic. The continual evolution of influenza viruses and emergence from animal reservoirs, leads to the need to quickly identify strains that pose a public health risk. In these studies, members of the National Institutes of Allergy and Infectious Diseases (NIAID) Centers for Excellence in Influenza Research and Surveillance (CEIRS) network worked together to demonstrate that the emerging canine H3 influenza viruses posed a low risk to public health and identified several therapeutic options in the event of an emergence. In addition to providing important new basic research, many lessons were learned that may be important in dealing with any emerging disease outbreak.
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- 2020
21. Maternally-Derived Antibodies Protect against Challenge with Highly Pathogenic Avian Influenza Virus of the H7N3 Subtype
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Daniel R. Perez, Adebimpe O. Obadan, Charles L. Hofacre, Zhimin Wan, Lucas M. Ferreri, Daniela S. Rajao, Ginger Geiger, Stivalis Cardenas-Garcia, and Silvia Carnaccini
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0301 basic medicine ,animal structures ,Immunology ,vaccination of chickens ,lcsh:Medicine ,Antibody level ,Biology ,Article ,03 medical and health sciences ,0302 clinical medicine ,Drug Discovery ,Pharmacology (medical) ,030212 general & internal medicine ,Viral shedding ,skin and connective tissue diseases ,Pharmacology ,Avian influenza virus ,Hemagglutination assay ,lcsh:R ,highly pathogenic avian influenza virus ,protection ,Virology ,Low pathogenic ,H7N3 ,Vaccination ,030104 developmental biology ,Infectious Diseases ,Highly Pathogenic Avian Influenza Virus ,embryonic structures ,maternally-derived antibodies ,biology.protein ,Antibody - Abstract
Vaccination of hens against influenza leads to the transfer of protective maternally-derived antibodies (MDA) to hatchlings. However, little is known about the transfer of H7N3 vaccine-induced MDA. Here, we evaluated transfer, duration, and protective effect of MDA in chickens against H7N3 HPAIV. To generate chickens with MDA (MDA (+)), 15-week-old White Leghorn hens were vaccinated and boosted twice with an inactivated H7N3 low pathogenic avian influenza virus vaccine, adjuvanted with Montanide ISA 71 VG. One week after the final boost, eggs were hatched. Eggs from non-vaccinated hens were hatched for chickens without MDA (MDA (&minus, )). Both MDA (+) and MDA (&minus, ) hatchlings were monitored weekly for antibody levels. Anti-HA MDA were detected by hemagglutination inhibition assay mostly until day 7 post-hatch. However, anti-nucleoprotein MDA were still detected three weeks post-hatch. Three weeks post-hatch, chickens were challenged with 106 EID50/bird of Mexican-origin H7N3 HPAIV. Interestingly, while 0% of the MDA (&minus, ) chickens survived the challenge, 95% of the MDA (+) chickens survived. Furthermore, virus shedding was significantly reduced by day 5 post-challenge in the MDA (+) group. In conclusion, MDA confers partial protection against mortality upon challenge with H7N3 HPAIV, as far as three weeks post-hatch, even in the absence of detectable anti-HA antibodies, and reduce virus shedding after challenge.
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- 2019
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22. Flexibility In Vitro of Amino Acid 226 in the Receptor-Binding Site of an H9 Subtype Influenza A Virus and Its Effect In Vivo on Virus Replication, Tropism, and Transmission
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Daniel R. Perez, Silvia Carnaccini, James C. Paulson, Daniela S. Rajao, Ginger Geiger, Adebimpe O. Obadan, Ana S. Gonzalez Reiche, Lucas M. Ferreri, Jefferson Santos, and Andrew J. Thompson
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receptor ,Hemagglutinin Glycoproteins, Influenza Virus ,Virus Replication ,medicine.disease_cause ,Madin Darby Canine Kidney Cells ,chemistry.chemical_compound ,Influenza A Virus, H9N2 Subtype ,Influenza A virus ,Amino Acids ,H9 subtype ,chemistry.chemical_classification ,0303 health sciences ,biology ,transmission ,Virus-Cell Interactions ,3. Good health ,Amino acid ,sialic acid ,Influenza Vaccines ,Leucine ,Protein Binding ,Immunology ,Hemagglutinin (influenza) ,Receptors, Cell Surface ,Quail ,Tropism ,Microbiology ,Virus ,Cell Line ,03 medical and health sciences ,Dogs ,Cell Line, Tumor ,Virology ,evolution ,Influenza, Human ,medicine ,Animals ,Humans ,avian viruses ,030304 developmental biology ,Binding Sites ,030306 microbiology ,zoonosis ,Sialic acid ,Glutamine ,HEK293 Cells ,Amino Acid Substitution ,chemistry ,Viral replication ,Influenza in Birds ,Insect Science ,biology.protein ,Chickens - Abstract
A single amino acid change at position 226 in the hemagglutinin (HA) from glutamine (Q) to leucine (L) has been shown to play a key role in receptor specificity switching in various influenza virus HA subtypes, including H9. We tested the flexibility of amino acid usage and determined the effects of such changes. The results reveal that amino acids other than L226 and Q226 are well tolerated and that some amino acids allow for the recognition of both avian and human influenza virus receptors in the absence of other changes. Our results can inform better avian influenza virus surveillance efforts as well as contribute to rational vaccine design and improve structural molecular dynamics algorithms., Influenza A viruses (IAVs) remain a significant public health threat, causing more than 300,000 hospitalizations in the United States during the 2015–2016 season alone. While only a few IAVs of avian origin have been associated with human infections, the ability of these viruses to cause zoonotic infections further increases the public health risk of influenza. Of these, H9N2 viruses in Asia are of particular importance as they have contributed internal gene segments to other emerging zoonotic IAVs. Notably, recent H9N2 viruses have acquired molecular markers that allow for a transition from avian-like to human-like terminal sialic acid (SA) receptor recognition via a single amino acid change at position 226 (H3 numbering), from glutamine (Q226) to leucine (L226), within the hemagglutinin (HA) receptor-binding site (RBS). We sought to determine the plasticity of amino acid 226 and the biological effects of alternative amino acids on variant viruses. We created a library of viruses with the potential of having any of the 20 amino acids at position 226 on a prototypic H9 HA subtype IAV. We isolated H9 viruses that carried naturally occurring amino acids, variants found in other subtypes, and variants not found in any subtype at position 226. Fitness studies in quails revealed that some natural amino acids conferred an in vivo replication advantage. This study shows the flexibility of position 226 of the HA of H9 influenza viruses and the resulting effect of single amino acid changes on the phenotype of variants in vivo and in vitro. IMPORTANCE A single amino acid change at position 226 in the hemagglutinin (HA) from glutamine (Q) to leucine (L) has been shown to play a key role in receptor specificity switching in various influenza virus HA subtypes, including H9. We tested the flexibility of amino acid usage and determined the effects of such changes. The results reveal that amino acids other than L226 and Q226 are well tolerated and that some amino acids allow for the recognition of both avian and human influenza virus receptors in the absence of other changes. Our results can inform better avian influenza virus surveillance efforts as well as contribute to rational vaccine design and improve structural molecular dynamics algorithms.
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- 2019
23. Plasticity of Amino Acid Residue 145 Near the Receptor Binding Site of H3 Swine Influenza A Viruses and Its Impact on Receptor Binding and Antibody Recognition
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Daniela S. Rajao, Daniel R. Perez, Nicola S Lewis, Ginger Geiger, James C. Paulson, Lucas M. Ferreri, Jefferson Santos, David F. Burke, Eugenio J. Abente, Adebimpe O. Obadan, Ana S. Gonzalez-Reiche, Andrew J. Thompson, Amy L. Vincent, Santos, Jefferson JS [0000-0001-5895-4163], Thompson, Andrew J [0000-0001-7865-1856], Perez, Daniel R [0000-0002-6569-5689], and Apollo - University of Cambridge Repository
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Models, Molecular ,Swine ,Immunology ,Hemagglutinin (influenza) ,Hemagglutinins, Viral ,Immunodominance ,Biology ,Antibodies, Viral ,Virus Replication ,Microbiology ,Antigenic drift ,Madin Darby Canine Kidney Cells ,03 medical and health sciences ,Residue (chemistry) ,chemistry.chemical_compound ,Dogs ,H3 subtype ,Polysaccharides ,Virology ,influenza vaccines ,Animals ,Humans ,Avidity ,hemagglutinin ,030304 developmental biology ,chemistry.chemical_classification ,virus evolution ,0303 health sciences ,swine influenza ,Binding Sites ,030306 microbiology ,Genetic Drift ,Amino acid ,Sialic acid ,HEK293 Cells ,Genetic Diversity and Evolution ,Biochemistry ,chemistry ,Amino Acid Substitution ,Influenza A virus ,Insect Science ,Viral evolution ,biology.protein ,Mutagenesis, Site-Directed ,Protein Binding - Abstract
The hemagglutinin (HA), a glycoprotein on the surface of influenza A virus (IAV), initiates the virus life cycle by binding to terminal sialic acid (SA) residues on host cells. The HA gradually accumulates amino acid substitutions that allow IAV to escape immunity through a mechanism known as antigenic drift. We recently confirmed that a small set of amino acid residues are largely responsible for driving antigenic drift in swine-origin H3 IAV. All identified residues are located adjacent to the HA receptor binding site (RBS), suggesting that substitutions associated with antigenic drift may also influence receptor binding. Among those substitutions, residue 145 was shown to be a major determinant of antigenic evolution. To determine whether there are functional constraints to substitutions near the RBS and their impact on receptor binding and antigenic properties, we carried out site-directed mutagenesis experiments at the single-amino-acid level. We generated a panel of viruses carrying substitutions at residue 145 representing all 20 amino acids. Despite limited amino acid usage in nature, most substitutions at residue 145 were well tolerated without having a major impact on virus replication in vitro. All substitution mutants retained receptor binding specificity, but the substitutions frequently led to decreased receptor binding. Glycan microarray analysis showed that substitutions at residue 145 modulate binding to a broad range of glycans. Furthermore, antigenic characterization identified specific substitutions at residue 145 that altered antibody recognition. This work provides a better understanding of the functional effects of amino acid substitutions near the RBS and the interplay between receptor binding and antigenic drift. IMPORTANCE The complex and continuous antigenic evolution of IAVs remains a major hurdle for vaccine selection and effective vaccination. On the hemagglutinin (HA) of the H3N2 IAVs, the amino acid substitution N 145 K causes significant antigenic changes. We show that amino acid 145 displays remarkable amino acid plasticity in vitro, tolerating multiple amino acid substitutions, many of which have not yet been observed in nature. Mutant viruses carrying substitutions at residue 145 showed no major impairment in virus replication in the presence of lower receptor binding avidity. However, their antigenic characterization confirmed the impact of the 145 K substitution in antibody immunodominance. We provide a better understanding of the functional effects of amino acid substitutions implicated in antigenic drift and its consequences for receptor binding and antigenicity. The mutation analyses presented in this report represent a significant data set to aid and test the ability of computational approaches to predict binding of glycans and in antigenic cartography analyses.
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- 2019
24. Avian Influenza Virus
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Lucas M. Ferreri, Jefferson Santos, Daniel R. Perez, Silvia Carnaccini, Stivalis Cardenas-Garcia, and Daniela S. Rajao
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Avian influenza virus ,Biology ,Virology - Published
- 2019
25. Development of a swine RNA polymerase I driven Influenza reverse genetics system for the rescue of type A and B Influenza viruses
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Daniel R. Perez, Matthew Angel, Troy C. Sutton, Stivalis Cardenas-Garcia, Daniela S. Rajao, Ginger Geiger, Ayush Kumar, Brittany Seibert, C. Joaquín Cáceres, and Lucas M. Ferreri
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0301 basic medicine ,Swine cells ,Swine ,viruses ,030106 microbiology ,Herpesvirus 1, Cercopithecine ,Virulence ,RNA polymerase II ,Article ,Virus ,03 medical and health sciences ,chemistry.chemical_compound ,RNA Polymerase I ,Virology ,RNA polymerase ,Influenza, Human ,RNA polymerase I ,Animals ,Humans ,Vector (molecular biology) ,biology ,Swine RNA polymerase I ,RNA ,Human cells ,Orthomyxoviridae ,Reverse Genetics ,Reverse genetics ,030104 developmental biology ,chemistry ,Influenza Vaccines ,biology.protein ,Influenza virus - Abstract
Influenza viruses are among the most significant pathogens of humans and animals. Reverse genetics allows for the study of molecular attributes that modulate virus host range, virulence and transmission. The most common reverse genetics methods use bi-directional vectors containing a host RNA polymerase (pol) I promoter to produce virus-like RNAs and a host RNA pol II promoter to direct the synthesis of viral proteins. Given the species-dependency of the pol I promoter and virus-host interactions that influence replication of animal-origin influenza viruses in human-derived cells, we explored the potential of using the swine RNA pol I promoter (spol1) in a bi-directional vector for rescuing type A and B influenza viruses (IAV and IBV, respectively) in swine and human cells. The spol1-based bi-directional plasmid vector led to efficient rescue of IAVs of different origins (human, swine, and avian) as well as IBV in both swine- and human-origin tissue culture cells. In addition, virus rescue was successful using a recombinant bacmid containing all eight segments of a swine origin IAV. In conclusion, the spol1-based reverse genetics system is a new platform to study influenza viruses and produce swine influenza vaccines with increased transfection efficiency.
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- 2021
26. Alternative Strategy for a Quadrivalent Live Attenuated Influenza Virus Vaccine
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Lucas M. Ferreri, Jefferson Santos, Zhimin Wan, Ginger Geiger, Stivalis Cardenas Garcia, Jing Liu, Daniel R. Perez, Daniela S. Rajao, and Silvia Carnaccini
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0301 basic medicine ,pandemic influenza ,viruses ,Immunology ,Biology ,universal vaccine platform ,Antibodies, Viral ,Vaccines, Attenuated ,Microbiology ,Proof of Concept Study ,Virus ,Madin Darby Canine Kidney Cells ,03 medical and health sciences ,Mice ,Viral Proteins ,Dogs ,Influenza A Virus, H1N1 Subtype ,protection efficiency ,Antigen ,influenza virus quadrivalent vaccine ,Virology ,Pandemic ,Vaccines and Antiviral Agents ,universal vaccine ,Live attenuated influenza vaccine ,Animals ,Humans ,influenza virus vaccines ,LAIV ,Hemagglutination assay ,Influenza A Virus, H3N2 Subtype ,virus diseases ,RNA-Dependent RNA Polymerase ,Immunoglobulin A ,Vaccination ,Titer ,Influenza B virus ,030104 developmental biology ,HEK293 Cells ,Immunization ,Influenza Vaccines ,Mice, Inbred DBA ,Insect Science ,Immunoglobulin G ,Mutation ,Female ,seasonal influenza - Abstract
Seasonal influenza viruses infect 1 billion people worldwide and are associated with ∼500,000 deaths annually. In addition, the never-ending emergence of zoonotic influenza viruses associated with lethal human infections and of pandemic concern calls for the development of better vaccines and/or vaccination strategies against influenza virus. Regardless of the strategy, novel influenza virus vaccines must aim at providing protection against both seasonal influenza A and B viruses. In this study, we tested an alternative quadrivalent live attenuated influenza virus vaccine (QIV) formulation whose individual components have been previously shown to provide protection. We demonstrate in proof-of principle studies in mice that the QIV provides effective protection against lethal challenge with either influenza A or B virus., Influenza virus infections continue to pose a major public health threat worldwide associated with seasonal epidemics and sporadic pandemics. Vaccination is considered the first line of defense against influenza. Live attenuated influenza virus vaccines (LAIVs) may provide superior responses compared to inactivated vaccines because the former can better elicit a combination of humoral and cellular responses by mimicking a natural infection. Unfortunately, during the 2013–2014, 2014–2015, and 2015–2016 seasons, concerns emerged about the effectiveness of the only LAIV approved in the United States that prevented the Advisory Committee on Immunization Practices (ACIP) from recommending its use. Such drawbacks open up the opportunity for alternative LAIV strategies that could overcome such concerns. Previously, we developed a combined strategy of temperature-sensitive mutations in the PB2 and PB1 segments and an epitope tag in the C terminus of PB1 that effectively attenuates influenza A viruses of avian and mammalian origin. More recently, we adopted a similar strategy for influenza B viruses. The resulting attenuated (att) influenza A and B viruses were safe, immunogenic, and protective against lethal influenza virus challenge in a variety of animal models. In this report, we provide evidence of the potential use of our att strategy in a quadrivalent LAIV (QIV) formulation carrying H3N2 and H1N1 influenza A virus subtype viruses and two antigenic lineages of influenza B viruses. In naive DBA/2J mice, two doses of the QIV elicited hemagglutination inhibition (HI) responses with HI titers of ≥40 and effectively protected against lethal challenge with prototypical pandemic H1N1 influenza A and influenza B virus strains. IMPORTANCE Seasonal influenza viruses infect 1 billion people worldwide and are associated with ∼500,000 deaths annually. In addition, the never-ending emergence of zoonotic influenza viruses associated with lethal human infections and of pandemic concern calls for the development of better vaccines and/or vaccination strategies against influenza virus. Regardless of the strategy, novel influenza virus vaccines must aim at providing protection against both seasonal influenza A and B viruses. In this study, we tested an alternative quadrivalent live attenuated influenza virus vaccine (QIV) formulation whose individual components have been previously shown to provide protection. We demonstrate in proof-of principle studies in mice that the QIV provides effective protection against lethal challenge with either influenza A or B virus.
- Published
- 2018
27. Development of an Alternative Modified Live Influenza B Virus Vaccine
- Author
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Isabel Aguirre, Adebimpe O. Obadan, Ginger Geiger, Zhimin Wan, Daniel R. Perez, Troy C. Sutton, Lucas M. Ferreri, Jefferson Santos, Courtney Finch, Ana S. Gonzalez-Reiche, and Diego Lopez
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0301 basic medicine ,T-Lymphocytes ,Immunology ,Genome, Viral ,antigenic variation ,Biology ,Vaccines, Attenuated ,Microbiology ,Virus ,Mice ,03 medical and health sciences ,Orthomyxoviridae Infections ,Immunity ,humoral immunity ,Virology ,Influenza, Human ,Vaccines and Antiviral Agents ,Antigenic variation ,Animals ,Humans ,Live attenuated influenza vaccine ,HA-tag ,Amino Acids ,Lung ,influenza B ,Duck embryo vaccine ,T cell immunity ,orthomyxovirus ,live vector vaccines ,Immunity, Humoral ,Vaccination ,Influenza B virus ,030104 developmental biology ,Vaccines, Inactivated ,Influenza Vaccines ,viral immunity ,Insect Science ,Mutation ,Humoral immunity - Abstract
Influenza B virus (IBV) is considered a major human pathogen, responsible for seasonal epidemics of acute respiratory illness. Two antigenically distinct IBV hemagglutinin (HA) lineages cocirculate worldwide with little cross-reactivity. Live attenuated influenza virus (LAIV) vaccines have been shown to provide better cross-protective immune responses than inactivated vaccines by eliciting local mucosal immunity and systemic B cell- and T cell-mediated memory responses. We have shown previously that incorporation of temperature-sensitive ( ts ) mutations into the PB1 and PB2 subunits along with a modified HA epitope tag in the C terminus of PB1 resulted in influenza A viruses (IAV) that are safe and effective as modified live attenuated ( att ) virus vaccines (IAV att ). We explored whether analogous mutations in the IBV polymerase subunits would result in a stable virus with an att phenotype. The PB1 subunit of the influenza B/Brisbane/60/2008 strain was used to incorporate ts mutations and a C-terminal HA tag. Such modifications resulted in a B/Bris att strain with ts characteristics in vitro and an att phenotype in vivo . Vaccination studies in mice showed that a single dose of the B/Bris att candidate stimulated sterilizing immunity against lethal homologous challenge and complete protection against heterologous challenge. These studies show the potential of an alternative LAIV platform for the development of IBV vaccines. IMPORTANCE A number of issues with regard to the effectiveness of the LAIV vaccine licensed in the United States (FluMist) have arisen over the past three seasons (2013–2014, 2014–2015, and 2015–2016). While the reasons for the limited robustness of the vaccine-elicited immune response remain controversial, this problem highlights the critical importance of continued investment in LAIV development and creates an opportunity to improve current strategies so as to develop more efficacious vaccines. Our laboratory has developed an alternative strategy, the incorporation of 2 amino acid mutations and a modified HA tag at the C terminus of PB1, which is sufficient to attenuate the IBV. As a LAIV, this novel vaccine provides complete protection against IBV strains. The availability of attenuated IAV and IBV backbones based on contemporary strains offers alternative platforms for the development of LAIVs that may overcome current limitations.
- Published
- 2017
28. Water Buffalos as carriers of Babesia bovis in Argentina
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Leonhard Schnittger, María Mesplet, Monica Florin-Christensen, Anabel Rodriguez, Daniel Benitez, Gustavo Asenzo, Mariana Dominguez, and Lucas M. Ferreri
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Veterinary medicine ,Buffaloes ,animal diseases ,Argentina ,Context (language use) ,Enzyme-Linked Immunosorbent Assay ,Biology ,General Biochemistry, Genetics and Molecular Biology ,History and Philosophy of Science ,parasitic diseases ,medicine ,Parasite hosting ,Animals ,Pathogen ,business.industry ,General Neuroscience ,Babesiosis ,Babesia bovis ,medicine.disease ,biology.organism_classification ,Bovine babesiosis ,Virology ,Livestock ,business ,Nested polymerase chain reaction ,geographic locations - Abstract
The tick-transmitted hemoprotozoan Babesia bovis is a major causative agent of bovine babesiosis, an often fatal disease of cattle. The disease is widespread in the northeastern region of Argentina, where an increasing part of the livestock is composed of water buffalos. Although clinical cases of buffalo babesiosis have not been reported so far, the pathogen-transmitting tick vector has been occasionally observed by us to be feeding on water buffalos. We therefore set out to examine whether buffalos may constitute a reservoir of the parasite. Competitive enzyme-linked immunosorbent assay (cELISA) detected B. bovis-specific antibodies in 20% of investigated buffalos (21/103), while direct detection of the pathogen by nested PCR was demonstrated in 34% of the animals (35/103). In one field, more than 60% of investigated animals (22/36) tested positive by nested PCR. These results are discussed in the context of buffalo babesiosis reported in other countries and in view of the currently effected control measures against bovine babesiosis in the region.
- Published
- 2009
29. Delivery of recombinant vaccines against bovine herpesvirus type 1 gD and Babesia bovis MSA-2c to mice using liposomes derived from egg yolk lipids
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Lucas M. Ferreri, Mariana Dominguez, Florencia Torrá, Monica Florin-Christensen, Patricia Ines Zamorano, Silvina Elizabeth Wilkowsky, and Anabel Rodriguez
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Male ,food.ingredient ,040301 veterinary sciences ,Protozoan Proteins ,Antibodies, Protozoan ,Antigens, Protozoan ,Antibodies, Viral ,Immunoglobulin G ,DNA vaccination ,0403 veterinary science ,Mice ,Viral Proteins ,03 medical and health sciences ,Plasmid ,food ,Babesiosis ,Yolk ,Animals ,Herpesvirus 1, Bovine ,030304 developmental biology ,Mice, Inbred BALB C ,0303 health sciences ,Liposome ,General Veterinary ,biology ,Membrane Proteins ,Viral Vaccines ,Babesia bovis ,Herpesviridae Infections ,04 agricultural and veterinary sciences ,biology.organism_classification ,Egg Yolk ,Virology ,Molecular biology ,3. Good health ,Naked DNA ,Liposomes ,biology.protein ,Female ,Animal Science and Zoology ,Antibody - Abstract
Liposomes prepared from total egg yolk lipid extracts were used to deliver experimental DNA vaccines to mice consisting of pCI-neo plasmids encoding bovine herpesvirus type 1 (BoHV-1) gD or Babesia bovis MSA-2c. A significantly higher proportion of mice in the B. bovis MSA-2c group, but not those in the BoHV-1 gD group, developed detectable immunoglobulin G responses when vaccinated with liposome encapsulated DNA in comparison with mice vaccinated with naked DNA. In both groups, antibody titres were similar between mice vaccinated with liposome encapsulated DNA and naked DNA.
- Published
- 2013
30. Influenza A virus reassortment in mammals gives rise to genetically distinct within-host subpopulations
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Ketaki Ganti, Anish Bagga, Silvia Carnaccini, Lucas M. Ferreri, Ginger Geiger, C. Joaquin Caceres, Brittany Seibert, Yonghai Li, Liping Wang, Taeyong Kwon, Yuhao Li, Igor Morozov, Wenjun Ma, Juergen A. Richt, Daniel R. Perez, Katia Koelle, and Anice C. Lowen
- Subjects
Science - Abstract
Virus reassortment drives genetic diversity and evolution and is governed by intra-host dynamics that are less well understood. Here, the authors characterise the within-host dynamics of influenza A virus reassortment in swine, ferrets and guinea pigs, considering their spatial distribution.
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- 2022
- Full Text
- View/download PDF
31. Improved detection of influenza A virus from blue‐winged teals by sequencing directly from swab material
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Lucas M. Ferreri, Lucia Ortiz, Ginger Geiger, Gonzalo P. Barriga, Rebecca Poulson, Ana Silvia Gonzalez‐Reiche, Jo Anne Crum, David Stallknecht, David Moran, Celia Cordon‐Rosales, Daniela Rajao, and Daniel R. Perez
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blue‐winged teal ,Guatemala ,Illumina ,influenza A virus ,next‐generation sequencing ,wild bird ,Ecology ,QH540-549.5 - Abstract
Abstract The greatest diversity of influenza A virus (IAV) is found in wild aquatic birds of the orders Anseriformes and Charadriiformes. In these birds, IAV replication occurs mostly in the intestinal tract. Fecal, cloacal, and/or tracheal swabs are typically collected and tested by real‐time RT‐PCR (rRT‐PCR) and/or by virus isolation in embryonated chicken eggs in order to determine the presence of IAV. Virus isolation may impose bottlenecks that select variant populations that are different from those circulating in nature, and such bottlenecks may result in artifactual representation of subtype diversity and/or underrepresented mixed infections. The advent of next‐generation sequencing (NGS) technologies provides an opportunity to explore to what extent IAV subtype diversity is affected by virus isolation in eggs. In the present work, we evaluated the advantage of sequencing by NGS directly from swab material of IAV rRT‐PCR‐positive swabs collected during the 2013–14 surveillance season in Guatemala and compared to results from NGS after virus isolation. The results highlight the benefit of sequencing IAV genomes directly from swabs to better understand subtype diversity and detection of alternative amino acid motifs that could otherwise escape detection using traditional methods of virus isolation. In addition, NGS sequencing data from swabs revealed reduced presence of defective interfering particles compared to virus isolates. We propose an alternative workflow in which original swab samples positive for IAV by rRT‐PCR are first subjected to NGS before attempting viral isolation. This approach should speed the processing of samples and better capture natural IAV diversity. OPEN RESEARCH BADGES This article has earned an Open Data Badge for making publicly available the digitally‐shareable data necessary to reproduce the reported results. The data is available at https://doi.org/10.5061/dryad.3h2n106.
- Published
- 2019
- Full Text
- View/download PDF
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