Recent Publications
2022
Shanley N Roach, Jessica K Fiege, Frances K Shepherd, Talia D Wiggen, Ryan C Hunter, Ryan A Langlois. Journal of Virology.
Influenza virus infection causes both respiratory and non-respiratory symptoms. Here, we characterize how respiratory influenza A virus infection alters the cellular environment in the intestine. We found that infection caused transient increases in changes in small intestine tuft cells, but had no impact on airway tuft cells. Type 1 and 2 innate lymphoid cells also increased in the small intestine during infection. In the absence of tuft cells, the increase in type 2 innate lymphoid cells was significantly blunted wherease type 1 cells were unaffected by the loss. Together, these results demonstrate tuft cells are necessary for infection-induced changes in type 2 innate lymphoid cells. These findings expand our understanding of the extra-pulmonary effects of respiratory virus infection as well as the role tuft cells play in regulating the intestinal environment.
Natural rodent model of viral transmission reveals biological features of virus population dynamics
Elizabeth J Fay, Keir M Balla, Shanley N Roach, Frances K Shepherd, Dira S Putri, Talia D Wiggen, Stephen A Goldstein, Mark J Pierson, Martin T Ferris, Claire E Thefaine, Andrew Tucker, Mark Salnikov, Valerie Cortez, Susan R Compton, Sergei V Kotenko, Ryan C Hunter, David Masopust, Nels C Elde, Ryan A Langlois
Emerging viruses threaten global health, but few experimental models can characterize the virus and host factors necessary for within- and cross-species transmission. Here, we exposed laboratory mice to pet store rodents - which harbor natural mouse pathogens - to study acute transmission through natural routes of exposure and infection. In this "dirty" mouse model, we identified various virus families that transmit to the exposed laboratory mice and describe: pathogen interactions that affect transmission, dynamics of transmission and dissemination within new hosts, and observation of cross-species transmission of a rat astrovirus. Together, this model system allows for the analysis of natural rodent virus transmission and is a platform to further characterize barriers to zoonosis.
2021
Genetic ancestry effects on the response to viral infection are pervasive but cell type specific
Haley E Randolph, Jessica K Fiege, Beth K ThieleN, Clayton K Mickelson, Mari Shiratori, João Barroso-Batista, Ryan A Langlois, Luis B Barreiro. Science.
Here we discuss findings from single-cell RNA sequencing of immune cells from individuals of European and African descent who were infected with influenza in vitro as it relates to different responses to infection, and how ancestry shapes those responses.
Mice with diverse microbial exposure histories as a model for preclinical vaccine testing
Jessica K Fiege, Katharine E Block, Mark J Pierson, Hezkiel Nanda, Frances K Shepherd, Clayton K Mickelson, J Michael Stolley, William E Matchett, Sathi Wijeyesinghe, David K Meyerholz, Vaiva Vezys, Steven S Shen, Sara E Hamilton, David Masopust, Ryan A Langlois. Cell Host Microbe.
We co-housed laboratory mice with pet-store mice, which have varied microbial exposures, and then assessed immune responses to influenza vaccines. Human transcriptional responses to influenza vaccination are better recapitulated in co-housed mice.
Intra- and Cross-Species Transmission of Astroviruses
Shanley N Roach and Ryan A Langlois. Viruses.
Here we discuss new research in astrovirus transmission
Nucleocapsid Vaccine Elicits Spike-Independent SARS-CoV-2 Protective Immunity
William E Matchett* , Vineet Joag, J Michael Stolley, Frances K Shepherd, Clare F Quarnstrom, Clayton K Mickelson, Sathi Wijeyesinghe, Andrew G Soerens , Samuel Becker , Joshua M Thiede 2, Eyob Weyu, Stephen D O'Flanagan, Jennifer A Walter, Michelle N Vu, Vineet D Menachery, Tyler D Bold, Vaiva Vezys, Marc K Jenkins, Ryan A Langlois, David Masopust. The Journal of Immunology. * contributed equally
Here we demonstrate that SARS-CoV-2 N-specific T cells can provide protection against challenge in hACE-2 transgenic mice and hamsters. These data support the use of conserved T cell antigen targets for future CoV vaccines.
Joshua M Thiede, Abigail R Gress, Samuel D Libby, Christine E Ronayne, William E Matchett, Brooke Noren, Joanne L Billings, Vineet D Menachery, Ryan A Langlois, Susan Kline, Tyler D Bold. Journal of Infectious Diseases.
We studied serum specimens from 19 hospitalized adults with COVID-19 randomized to treatment with remdesivir or placebo. We assessed severe acute respiratory syndrome coronavirus 2 antibody responses and identified cytokine signatures, using hierarchical clustering. We identified no clear immunologic trends attributable to remdesivir treatment.
Segment-specific kinetics of mRNA, cRNA and vRNA accumulation during influenza infection
Thu Phan, Elizabeth J Fay, Zion Lee, Stephanie Aron, Wei-Shou Hu, Ryan A Langlois. Journal of Virology.
We developed a new approach that allows the methodical study of IAV replication kinetics, shedding light on many interesting features of IAV replication biology. This study advances our understanding of the kinetics of IAV replication and will help to facilitate future research in the field.
Jessica K Fiege, Joshua M Thiede, Hezkiel Arya Nanda, William E Matchett, Patrick J Moore, Noe Rico Montanari, Beth K Thielen, Jerry Daniel, Emma Stanley, Ryan C Hunter, Vineet D Menachery, Steven S Shen, Tyler D Bold, Ryan A Langlois. PLOS PATHOGENS.
We used flow cytometry and single cell RNA-sequencing to understand how the heterogeneity of this diverse cell population contributes to elements of viral tropism and pathogenesis, antiviral immunity, and treatment response to remdesivir. We found that ciliated cells are the predominant cell target of SARS-CoV-2, and that heavily infected secretory cells expressed abundant IL-6, a potential mediator of COVID-19 pathogenesis.
Mouse SARS-Cov-2 Epitope Reveals Infection and Vaccine-Elicited CD8 T Cell Responses
Vineet Joag, Sathi Wijeyesinghe, J Michael Stolley, Clare F Quarnstrom, Thamotharampillai Dileepan, Andrew G Soerens, Jules A Sangala, Stephen D O'Flanagan, Noah V Gavil, Sung-Wook Hong, Siddheshvar Bhela, Sailaja Gangadhara, Eyob Weyu, William E Matchett, Joshua Thiede, Venkatramana Krishna, Maxim C-J Cheeran, Tyler D Bold, Rama Amara, Peter Southern, Geoffrey T Hart, Luca Schifanella, Vaiva Vezys, Marc K Jenkins, Ryan A Langlois, David Masopust. The Jounral of Immunology.
Whereas many COVID-19 vaccines focus on establishing humoral immunity to viral spike protein, vaccine-elicited T cell immunity may bolster durable protection or cross-reactivity with viral variants. To better enable mechanistic and vaccination studies in mice, we identified a dominant CD8 T cell SARS-CoV-2 nucleoprotein epitope. In mice, i.m. prime-boost nucleoprotein vaccination with heterologous vectors favored systemic CD8 T cell responses, whereas intranasal boosting favored respiratory immunity.
2020
Cell type- and replication stage-specific influenza virus responses in vivo
Elizabeth J Fay, Stephanie L Aron, Marissa G Macchietto, Matthew W Markman, Katharina Esser-Nobis, Michael Gale Jr, Steven Shen, Ryan A Langlois. PLos Pathogens.
Activation of the innate immune response is important for controlling early virus replication and spread. Here, we use single-cycle influenza A viruses to characterize the early immune response. We found that the magnitude of virus replication contributes to antiviral gene expression within infected cells prior to the induction of a global response. These data offer insight into the early mechanisms of antiviral gene activation during influenza A infection.
Thomas J Gniadek, Joshua M Thiede, William E Matchett, Abigail R Gress, Kathryn A Pape, Marc K Jenkins, Vineet D Menachery, Ryan A Langlois, Tyler D Bold. The Jounral of AABB - Transfusion.
We determined the antigen binding activity of convalescent plasma units from 47 individuals with a history of non-severe COVID-19 using three clinical diagnostic serology assays with different SARS-CoV-2 targets. We compared these results with functional neutralization activity using a fluorescent reporter strain of SARS-CoV-2 in a microwell assay. Donors age 48-75 had the highest neutralization activity.
Nelson AC, Auch B, Schomaker M, Gohl DM, Grady P, Johnson D, Kincaid R, Karnuth KE, Daniel J, Fiege JK, Fay EJ, Bold TD, Langlois RA, Beckman KB, and Yohe S. BioRxiv.
The COVID-19 global pandemic is an unprecedented health emergency. We rapidly developed a qRT-PCR SARS-CoV-2 detection assay using a 384-well format and tested its analytic performance across multiple nucleic acid extraction kits. Our methods and results provide valuable information for other high-complexity laboratories seeking to develop effective, local, laboratory-developed procedures with high-throughput capability to detect SARS-CoV-2.
Virus-induced transposable element expression up-regulation in human and mouse host cells.
Macchietto MG, Langlois RA, and Shen S. Life Science Alliance.
By reanalyzing sequencing data sets from dozens of virus infection studies, we found that genome-wide transposon expression up-regulation in host cells occurs near antiviral response genes and exists in all studies regardless of virus, species, and host cell tissue, indicating that transposon up-regulation is a common phenomenon during virus infection.
2019
Long-term surviving influenza infected cells evade CD8+ T cell mediated clearance.
Fiege JK, Stone IA, Dumm RE, Waring BM, Fife BT, Agudo J, Brown B, Heaton NS, and Langlois RA. PLoS Pathogens.
Using a virus expressing Cre recombinase to permanently label infected cells in a Cre-inducible reporter mouse, we previously discovered infected club cells that survive both lytic virus replication and CD8+ T cell-mediated clearance. We demonstrate that ciliated epithelial cells, type I and type II alveolar cells can also become stable in the lung long-term and demonstrate enhanced proliferation compared to uninfected cells.
Hemann EA, Green R, Turnbull JB, Langlois RA, Savan R, and Gale Jr M. Nature Immunology.
Type III interferon (IFN-λ) is important for innate immune protection at mucosal surfaces and has therapeutic benefit against influenza A virus (IAV) infection. However, the mechanisms by which IFN-λ programs adaptive immune protection against IAV are undefined. Here we found that IFN-λ signaling in dendritic cell (DC) populations was critical for the development of protective IAV-specific CD8+ T cell responses.
Fiege JK, Stone IA, Fay EJ, Markman M, Wijeyesinghe S, Macchietto MG, Shen S, Masopust D and Langlois RA. Journal of Immunology.
Resident memory T cells (TRM) in the lung are vital for heterologous protection against influenza A virus (IAV). Environmental factors are necessary to establish lung TRM; however, the role of T cell-intrinsic factors like TCR signal strength have not been elucidated. In this study, we investigated the impact of TCR signal strength on the generation and maintenance of lung TRM after IAV infection.
TCR Affinity Biases Th Cell Differentiation by Regulating CD25, Eef1e1, and Gbp2.
Kotov DI, Mitchell JS, Pengo T, Rueld C, Way SS, Langlois RA, Fife BT and Jenkins MK. Journal of Immunology.
The affinity of the TCR interaction withmicrobial peptide:MHC class II plays a role in Th differentiation by mechanisms that are not completely understood. We found that low-affinity TCRs biased mouse naive T cells to become T follicular helper cells, whereas higher-affinity TCRs promoted the formation of Th1 or Th17 cells. These results suggest that TCR affinity regulates genes in naive T cells that control the differentiation process.
Hornick EE, Zacharias ZR, Miller AM, Langlois RA, Chen P, Legge KL, Bishop GA, Cassel SL, and Sutterwala FS. Journal of Clinical Investigation.
Generation and maintenance of robust IAV-specific T cell responses relies on T cell interactions with dendritic cells. In this study, we explore the role of nucleotide-binding domain leucine-rich repeat containing receptor family member NLRC4 in modulating the DC phenotype during IAV infection. Our data support a novel role for NLRC4 in regulating the phenotype of lung DCs during a respiratory viral infection
Non-lytic clearance of influenza B virus from infected cells preserves epithelial barrier function.
Dumm RE, Fiege JK, Waring BM, Langlois RA and Heaton NS. Nature Communications.
Influenza B virus (IBV) is an acute, respiratory RNA virus that has been assumed to induce the eventual death of all infected cells. We and others have shown however, that infection with apparently cytopathic viruses does not necessarily lead to cell death; some cells can intrinsically clear the virus and persist in the host long-term.
Engineered Small-Molecule Control of Influenza A Virus Replication.
Fay EJ, Aron SL, Stone IA, Waring BM, Plemper RL and Langlois RA. Journal of Virology.
New influenza A virus vaccines must be tested in challenge studies, posing significant safety problems, since current pharmacological interventions for IAV are poorly efficacious. We have generated a virus expressing a small-molecule-assisted shutoff (SMASh) tag as a safety switch for IAV replication.
2018
Sjaastad LE, Fay EJ, Fiege JK, Macchietto MG, Stone IA, Markman MW, Shen S, and Langlois RA. PNAS.
Influenza virus has a broad cellular tropism in the respiratory tract. Infected epithelial cells sense the infection and initiate an antiviral response. To define the antiviral response at the earliest stages of infection we used a series of single-cycle reporter viruses. These viral probes demonstrated cells in vivo harbor a range in magnitude of virus replication.
MicroRNA-Attenuated Virus Vaccines.
Fay EJ and Langlois RA. Non-Coding RNA.
Live-attenuated vaccines are the most effective way to establish robust, long-lasting immunity against viruses. However, the possibility of reversion to wild type replication and pathogenicity raises concerns. The use of host-derived microRNAs to attenuate viruses has been accomplished in an array of biological contexts. The broad assortment of effective tissue- and species-specific miRNAs, and the ability to target a virus with multiple miRNAs, allow for targeting to be tailored to the virus.
Memory CD8 T cells mediate severe immunopathology following respiratory syncytial virus infection.
Stoley M, Knudson C, Hartwig S, Pewe L, Meyerholz D, Langlois RA, Harty JT, Varga SM. PLoS Pathogens.
Memory T cells can provide protection from re-infection by respiratory viruses such as influenza and SARS. However, the relative contribution of memory CD8 T cells in providing protection against respiratory syncytial virus (RSV) infection is currently unclear.
2017
microRNA based attenuation of influenza virus across susceptible hosts.
Waring BM, Sjaastad LE, Fiege JK, Fay EJ, Reyes I, Moriarity B, Langlois RA. Journal of Virology.
Influenza A virus drives significant morbidity and mortality in humans and livestock. Annual circulation of the virus in livestock and waterfowl contributes to severe economic disruption and increases the risk of zoonotic transmission of novel strains into the human population, where there is no preexisting immunity.
2016
Club cells surviving influenza A virus infection induce temporal nonspecific antiviral immunity.
Hamilton JR, Sachs D, Lim JK, Langlois RA, Palese P, Heaton NH. PNAS.
A brief window of antigen-nonspecific protection has been observed after influenza A virus (IAV) infection. Although this temporary immunity has been assumed to be the result of residual nonspecific inflammation, this period of induced immunity has not been fully studied.
2015
Hemann EA, Sjaastad LE, Langlois RA, Legge KL. Journal of Virology.
Following influenza A virus (IAV) infection, development of a robust IAV-specific CD8 T cell response is required for clearance of primary infection and enhances memory protection. Following IAV infection, plasmacytoid dendritic cells (pDC) or CD8α(+) DC regulate pulmonary effector CD8 T cell responses within the lung.
Investigating Influenza A Virus infection: Tools to track infection and limit tropism.
Fiege JK, Langlois RA. Journal of Virology.
Influenza A viruses display a broad cellular tropism within the respiratory tracts of mammalian hosts. Uncovering the relationship between tropism and virus immunity, pathogenesis, and transmission will be critical for the development of therapeutic interventions. Here we discuss recent developments of several recombinant strains of influenza A virus.
2014
Long-term survival of influenza virus infected club cells drives immunopathology.
Heaton NH, Langlois RA, Sachs D, Lim JK, Palese P, tenOever BR. Journal of Experimental Medicine.
Respiratory infection of influenza A virus (IAV) is frequently characterized by extensive immunopathology and proinflammatory signaling that can persist after virus clearance. In this report, we identify cells that become infected, but survive, acute influenza virus infection.
The mammalian response to RNA virus infections is independent of RNAi.
Backes S, Langlois RA, Schmid S, Shim JV, Sachs D, tenOever BR. Cell Report.
A successful cellular response to virus infection is essential for evolutionary survival. In plants, arthropods, and nematodes, cellular antiviral defenses rely on RNAi. Interestingly, the mammalian response to virus is predominantly orchestrated through interferon (IFN)-mediated induction of antiviral proteins.
2013
MicroRNA-based strategy to mitigate the risk of gain-of-function influenza studies.
Langlois RA, Albrecht RA, Kimble B, Sutton T, Shapiro JS, Finch C, Angel M, Chua MA, Gonzalez-Reiche AS, Xu K, PerezD, Garcia-Sastre A, tenOever BR. Nature Biotechnology.
Recent gain-of-function studies in influenza A virus H5N1 strains revealed that as few as three-amino-acid changes in the hemagglutinin protein confer the capacity for viral transmission between ferrets. As transmission between ferrets is considered a surrogate indicator of transmissibility between humans, these studies raised concerns about the risks of gain-of-function influenza A virus research.
Influenza A Virus Utilizes Suboptimal Splicing to Coordinate the Timing of Infection.
Chua MA, Perez JT, Schmid S, Langlois RA, tenOever BR. PNAS.
Influenza A virus is unique as an RNA virus in that it replicates in the nucleus and undergoes splicing. With only ten major proteins, the virus must gain nuclear access, replicate, assemble progeny virions in the cytoplasm, and then egress. We find that utilization of an erroneous splice site ensures the slow accumulation of the viral nuclear export protein (NEP) while generating excessive levels of an antagonist that inhibits the cellular response to infection.
2012
Pica N, Langlois RA, Krammer F, Margarine I, and Palese, P. Journal of Virology.
Immunological changes associated with age contribute to the high rates of influenza virus morbidity and mortality in the elderly. Compounding this problem, aged individuals do not respond to vaccination as well as younger, healthy adults. Using a live influenza virus with a truncated nonstructural protein 1 (NS1), we are able to stimulate cellular and humoral immune responses of aged mice comparable to levels seen in young mice.
Langlois RA, Varble A, Chua MA, García-Sastre A, tenOever BR. PNAS.
A coordinated innate and adaptive immune response, orchestrated by antigen presenting cells, is required for effective clearance of influenza A virus. Although IAV primarily infects epithelial cells of the upper respiratory tract, APCs are also susceptible. We demonstrate that inhibiting IAV replication in APCs in vivo did not alter clearance, or the generation of IAV-specific CD8 T cells, suggesting that cross-presentation is sufficient for T lymphocyte activation.
Evidence for a cytoplasmic microprocessor of pri-miRNAs.
Shapiro JS, Langlois RA, Pham AM, tenOever BR. RNA.
While miRNA biogenesis occurs in a stepwise fashion, initiated by the nuclear microprocessor, rare non-canonical miRNAs have also been identified. Here, we characterize the molecular components and unique attributes associated with the processing of virus-derived cytoplasmic primary miRNAs. This research describes a unique function for Drosha in the processing of highly structured cytoplasmic RNAs in the context of virus infection.
Replication in cells of hematopoietic origin is necessary for Dengue virus dissemination.
Pham AM, Langlois RA, tenOever BR. PLoS Pathogens.
Dengue virus (DENV) is a mosquito-borne pathogen for which no vaccine or specific therapeutic is available. We exploited hematopoietic-specific microRNA-142 (miR-142) to control virus tropism by inserting tandem target sites into the virus to restrict replication exclusively in this cell population. This suggests that these immune cells are the predominant sources of virus amplification.
In vivo delivery of cytoplasmic RNA virus-derived miRNAs.
Langlois RA, Shapiro JS, Pham AM, tenOever BR. Molecular Therapy.
The discovery of microRNAs has presented an attractive strategy for therapeutics that is currently limited by in vivo delivery constraints. We describe the generation of a single-stranded, cytoplasmic virus of negative polarity capable of producing functional miRNAs within a wide range of tissues, and sustaining prolonged expression capable of achieving measurable knockdown of physiological targets in vivo.