Biology subjects

Geiger, G.

Publications and source records attributed to Geiger, G..

12 recordsLinked to original sources

bioRxiv2026

Contemporary Victoria and Yamagata Influenza B Viruses Elicit Lineage-Specific Differences in Innate Immunity

The two influenza B virus (FLUBV) lineages, Victoria and Yamagata, have continued to diverge since their separation in the 1970s, resulting in distinct antigenic characteristics and differences in immunity and cross-protection. Human epidemiological observations and experimental studies have identified lineage-specific differences in adaptive immune responses elicited by contemporary viruses, suggesting that innate immune signaling may contribute to these divergent outcomes. To understand in-depth immunity to contemporary influenza B viruses, we compared viral replication, cytokine production, gene expression, and cellular tropism following infection with representative Victoria- and Yamagata-lineage viruses. We found that, despite broadly similar cytokine profiles in the ferret upper respiratory tract, the two lineages exhibited distinct innate immune kinetics. Yamagata viruses induced rapid early expression of antiviral and inflammatory genes, including SOCS1 and multiple interferon-stimulated genes, whereas Victoria viruses displayed delayed innate immune activation accompanied by greater viral replication. Consistent with the ferret data, Yamagata viruses also induced elevated SOCS1 expression in human PBMCs early after infection. In addition, Yamagata viruses exhibited broader cellular tropism, infecting a wider range of immune cell populations than Victoria viruses. Together, these findings demonstrate that contemporary influenza B virus lineages distinctly engage the host immune system and provide new insight into how lineage-specific innate immune responses may contribute to differences in immunity, cross-protection, and the divergent evolutionary trajectories of Victoria and Yamagata viruses

microbiology↗

bioRxiv2025

Passage of human-origin influenza A virus in swine tracheal epithelial cells selects for adaptive mutations in the hemagglutinin gene

AbstractFrequent spillover of influenza A viruses from humans to swine contributes to the increasing diversity of influenza viruses circulating in pigs. Although these events are common, little is known about the adaptation processes that take place when viruses jump between the two species. We examined the changes that occurred during serial passages of a reassortant H3N2 virus (VIC11pTRIG) containing human seasonal surface genes (Hemagglutinin and Neuraminidase) and a swine-adapted internal gene constellation in differentiated primary swine tracheal epithelial cells (pSTECs). The VIC11pTRIG reassortant virus was serially passaged 8 times in pSTECs and compared to a control swine-adapted strain (OH/04p) containing the same internal gene constellation. Viral RNA from passages 0 (inoculum), 1, 3, 4-8 were sequenced via next generation or Sanger sequencing. Hemagglutinin diversity was highest at passage 3. Two amino acid mutations in the Hemagglutinin protein (N165K and N216K) were fixed at passages 7 and 5, respectively. These changes were associated with increased fitness of the virus in pSTECs compared to the original parental strain. Our results suggest that the adaptation of human seasonal H3N2 to swine cells may lead to the selection of HA mutations located near the receptor binding site. These mutations may result in increased fitness of human-origin H3N2 strains to adapt in swine.

microbiology↗

bioRxiv2024

Differential Cross-Protective Immunity is Elicited by Infection with Contemporary Influenza B Lineage Viruses

Influenza B virus (FLUBV) significantly contributes to the influenza disease burden and has complicated vaccine development and efficacy, yet remains understudied compared to its counterpart, influenza A virus (FLUAV). Since its isolation in 1940, FLUBV has diverged into two antigenically distinct lineages: Victoria (B/Vic) and Yamagata (B/Yam). Recent human studies and epidemiological modeling reveal differences in immunity elicited by each FLUBV lineage, contributing to higher reinfection rates following B/Yam infection. To investigate disparities in FLUBV lineage cross-protection and immunity, we examined the effects of lineage-specific prior immunity on FLUBV reinfection dynamics. Mice were infected with representative B/Vic and B/Yam viruses from evolutionary distinct clades and subsequently reinfected with heterolineal viruses (i.e., B/Vic [->] B/Yam and B/Yam [->] B/Vic) to assess the extent of protection elicited between the lineages. Using this validated challenge model, we explored potential mechanisms underlying the asymmetric reinfection dynamics observed between the lineages. Our findings align with human observations, indicating that contemporary B/Vic viruses confer cross-protection against contemporary B/Yam infections, whereas contemporary B/Yam viruses do not provide the same degree of protection. Furthermore, we demonstrated that serum antibodies elicited by hemagglutinin vaccination cannot account for the observed heterolineal protection. Rather, antibodies targeting the viral neuraminidase (NA) may play a significant role in eliciting cross-protection to subsequent FLUBV infection. Our findings define asymmetric cross-protection resulting from contemporary FLUBV infection and suggest NA as a potential significant contributor to heterolineal FLUBV protection. This asymmetric immunity may also help explain the proposed extinction of B/Yam viruses since the COVID-19 pandemic. ImportanceInfluenza B viruses (FLUBV) consist of two divergently evolving lineages, Victoria (B/Vic) and Yamagata (B/Yam). Contemporary isolates from these lineages exhibit increased endemic activity and higher evolutionary rates while utilizing distinct mechanisms for evolutionary success. This is exemplified by novel seasonal infection dynamics with Influenza A viruses, differences in cross-protection elicited between the FLUBV lineages, and the potential extinction of B/Yam following the COVID-19 pandemic. We explore FLUBV infection dynamics utilizing contemporary viruses to define the asymmetric immunity elicited between the lineages. Contemporary Yamagata viruses are unable to confer the same breadth of protection as Victoria viruses. This may help explain the higher reinfection rates for Yamagata viruses and suggest a potential contributor to the extinction of this lineage.

microbiology↗

bioRxiv2024

FLUAV RAM-IGIP: A modified live influenza virus vaccine that enhances humoral and mucosal responses against influenza

Current influenza A vaccines fall short, leaving both humans and animals vulnerable. To address this issue, we have developed attenuated modified live virus (MLV) vaccines against influenza using genome rearrangement techniques targeting the internal gene segments of FLUAV. The rearranged M2 (RAM) strategy involves cloning the M2 ORF downstream of the PB1 ORF in segment 2 and incorporating multiple early stop codons within the M2 ORF in segment 7. Additionally, the IgA-inducing protein (IGIP) coding region was inserted into the HA segment to further attenuate the virus and enhance protective mucosal responses. RAM-IGIP viruses exhibit similar growth rates to wild type (WT) viruses in vitro and remain stable during multiple passages in cells and embryonated eggs. The safety, immunogenicity, and protective efficacy of the RAM-IGIP MLV vaccine against the prototypical 2009 pandemic H1N1 strain A/California/04/2009 (H1N1) (Ca/04) were evaluated in Balb/c mice and compared to a prototypic cold-adapted live attenuated virus vaccine. The results demonstrate that the RAM-IGIP virus exhibits attenuated virulence in vivo. Mice vaccinated with RAM-IGIP and subsequently challenged with an aggressive lethal dose of the Ca/04 strain exhibited complete protection. Analysis of the humoral immune response revealed that the inclusion of IGIP enhanced the production of neutralizing antibodies and augmented the antibody-dependent cellular cytotoxicity response. Similarly, the RAM-IGIP potentiated the mucosal immune response against various FLUAV subtypes. Moreover, increased antibodies against NP and NA responses were observed. These findings support the development of MLVs utilizing genome rearrangement strategies in conjunction with the incorporation of immunomodulators. IMPORTANCECurrent influenza vaccines offer suboptimal protection, leaving both humans and animals vulnerable. Our novel attenuated MLV vaccine, built by rearranging FLUAV genome segments and incorporating the IgA-inducing protein, shows promising results. This RAM-IGIP vaccine exhibits safe attenuation, robust immune responses, and complete protection against lethal viral challenge in mice. Its ability to stimulate broad-spectrum humoral and mucosal immunity against diverse FLUAV subtypes makes it a highly promising candidate for improved influenza vaccines.

immunology↗

bioRxiv2023

Antigenic mapping of the hemagglutinin of the H9 subtype influenza A viruses using sera from Japanese quail (Coturnix c. japonica).

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.

microbiology↗

bioRxiv2022

Independent evolution of influenza A virus H1N1 in pigs in Guatemala

Commercial swine farms provide unique systems for interspecies transmission of influenza A viruses (FLUAVs) at the animal-human interface. Bidirectional transmission of FLUAVs between pigs and humans plays a significant role in the generation of novel strains that become established in the new host population. Active FLUAV surveillance was conducted for two years on a commercial pig farm in Southern Guatemala with no history of FLUAV vaccination. Nasal swabs (n=2,094) from fattening pigs (6 to 24 weeks old) with respiratory signs were collected from May 2016 to February 2018. Swabs were screened for FLUAV by RRT-PCR and full virus genomes of FLUAV-positive swabs were sequenced by next-generation sequencing (NGS). FLUAV prevalence was 12.0% (95% CI: 10.6% - 13.4%) with two distinct periods of high infection. All samples were identified as FLUAVs of the H1N1 subtype within the H1 swine clade 1A.3.3.2 and whose ancestors are the human origin 2009 H1N1 influenza pandemic virus (H1N1 pdm09). Compared to the prototypic reference segment sequence, 10 amino acid signatures were observed on relevant antigenic sites on the hemagglutinin. We also found that Guatemalan swine-origin FLUAVs show independent evolution from other H1N1 pdm09 FLUAVs circulating in Central America. The zoonotic risk of these viruses remains unknown, but strongly calls for continued FLUAV surveillance in pigs in Guatemala. ImportanceDespite increased surveillance efforts, the epidemiology of FLUAVs circulating in swine in Latin America remains understudied. For instance, the 2009 H1N1 influenza pandemic strain (H1N1 pdm09) emerged in Mexico, but its circulation remained undetected in pigs. In Central America, Guatemala is the country with the largest swine industry. We found a unique group of H1N1 pdm09 sequences that suggests independent evolution from similar viruses circulating in Central America. These viruses may represent the establishment of a novel genetic lineage with the potential to reassort with other co-circulating viruses, and whose zoonotic risk remains to be determined.

evolutionary biology↗

bioRxiv2022

Epistasis reduces fitness costs of influenza A virus escape from stem-binding antibodies

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.

microbiology↗

bioRxiv2022

Pathobiology and dysbiosis of the respiratory and intestinal microbiota in 14 months old Golden Syrian hamsters infected with SARS-CoV-2

The pandemic of severe acute respiratory syndrome coronavirus 2 (SARS2) affected the geriatric population. Among research models, Golden Syrian hamsters (GSH) are one of the most representative to study SARS2 pathogenesis and host responses. However, animal studies that recapitulate the effects of SARS2 in the human geriatric population are lacking. To address this gap, we inoculated 14 months old GSH (resembling people over 60 years old) with a prototypic ancestral strain of SARS2 and studied the effects on virus pathogenesis, virus shedding, and respiratory and gastrointestinal microbiome changes. SARS2 infection led to high vRNA loads in the nasal turbinates (NT), lungs, and trachea as well as higher pulmonary lesions scores later in infection. Dysbiosis throughout SARS2 disease progression was observed in the pulmonary microbial dynamics with the enrichment of opportunistic pathogens (Haemophilus, Fusobacterium, Streptococcus, Campylobacter, and Johnsonella) and microbes associated with inflammation (Prevotella). Changes in the gut microbial community also reflected an increase in multiple genera previously associated with intestinal inflammation and disease (Helicobacter, Mucispirillum, Streptococcus, unclassified Erysipelotrichaceae, and Spirochaetaceae). Influenza A virus (FLUAV) pre-exposure resulted in slightly more pronounced pathology in the NT and lungs early on (3 dpc), and more notable changes in lungs compared to the gut microbiome dynamics. Similarities among aged GSH and the microbiome in critically ill COVID-19 patients, particularly in the lower respiratory tract, suggest that GSHs are a representative model to investigate microbial changes during SARS2 infection. The relationship between the residential microbiome and other confounding factors, such as SARS2 infection, in a widely used animal model, contributes to a better understanding of the complexities associated with the host responses during viral infections. Author SummaryThe SARS-CoV-2 pandemic led to millions of human losses, notably affecting the geriatric population, who are at greater risk of developing acute respiratory distress infection leading to prolonged hospitalization and death. However, the mechanism of age-related pathogenicity is not fully understood. Here, we utilized an aged Syrian hamster model resembling ~60-year-old humans to analyze the pathobiology, host response, and effects of SARS2 on the respiratory and intestinal microbiome. We identified specific microbial markers observed in severe COVID-19 patients within the lungs of aged hamsters infected with SARS-CoV-2. Prior influenza A virus (H1N1) exposure amplified these changes. Similarities among aged GSH and critically ill COVID-19 patients suggest that GSHs are a valuable model for investigating microbial changes during SARS2 infection. The relationship between the age, residential microbiome and viral pathogens contributes to a better understanding of the complexities associated with the host responses during viral infection while limiting potential environmental factors that may contribute to inter-individual variation.

microbiology↗

bioRxiv2022

Transmission of human influenza A virus in pigs selects for adaptive mutations on the HA gene

Influenza A viruses (IAV) cause respiratory diseases in many host species, including humans and pigs. The spillover of IAV 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 IAVs 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 IAV to transmit and become established in pigs. We generated a reassortant IAV containing surface gene segments from a human IAV strain and internal gene segments from the 2009 pandemic and TRIG IAV lineages and demonstrated that it can replicate and transmit in pigs. Sequencing and variant calling analysis identified a mutant that emerged during replication in pigs, which was mapped to a region near the receptor binding site of the hemagglutinin (HA). The variant was present in all contact pigs and replicated more efficiently in differentiated swine tracheal cells compared to the virus containing the wildtype human-origin HA. 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. ImportanceInfluenza A viruses (IAV) cause respiratory disease in several species, including humans and pigs. The bidirectional transmission of IAV 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 IAV to become established in pigs. In this study, we generated reassortant viruses containing human seasonal HA and 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 mutation 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.

microbiology↗

bioRxiv2022

Robustness of the ferret model for influenza risk assessment studies: a cross-laboratory exercise

Ferrets represent the preferred animal model for assessing the transmission potential of newly emerged zoonotic influenza viruses. However, heterogeneity among established experimental protocols and facilities across different laboratories may lead to variable results, complicating interpretation of transmission experimental data. Between 2018-2020, a global exercise was conducted by 11 participating laboratories to assess the range of variation in ferret transmission experiments using two common stock H1N1 influenza viruses that possess different transmission characteristics in ferrets. Inoculation route, dose, and volume were standardized, and all participating laboratories followed the same experimental conditions for respiratory droplet transmission, including a strict 1:1 donor:contact ratio. Additional host and environmental parameters likely to affect influenza transmission kinetics were monitored throughout. Overall transmission outcomes for both viruses across 11 laboratories were concordant, suggesting the robustness of the ferret model for zoonotic influenza risk assessment. To attain high confidence in identifying zoonotic influenza viruses with moderate-to-high or low transmissibility, our analyses support that as few as three but as many as five laboratories, respectively, would need to independently perform viral transmission experiments with concordant results. This exercise facilitates the development of a more homogenous protocol for ferret transmission experiments that are employed for the purposes of risk assessment.

microbiology↗

bioRxiv2022

Influenza A virus reassortment in mammals gives rise to genetically distinct within-host sub-populations

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.

microbiology↗

bioRxiv2021

Mild and severe SARS-CoV-2 infection induces respiratory and intestinal microbiome changes in the K18-hACE2 transgenic mouse model

Transmission of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has resulted in millions of deaths and declining economies around the world. K18-hACE2 mice develop disease resembling severe SARS-CoV-2 infection in a virus dose-dependent manner. The relationship between SARS-CoV-2 and the intestinal or respiratory microbiome is not fully understood. In this context, we characterized the cecal and lung microbiome of SARS-CoV-2 challenged K18-hACE2 transgenic mice in the presence or absence of treatment with the Mpro inhibitor GC376. Cecum microbiome showed decreased Shannon and Inv Simpson diversity index correlating with SARS-CoV-2 infection dosage and a difference of Bray-Curtis dissimilarity distances among control and infected mice. Bacterial phyla such as Firmicutes, particularly Lachnospiraceae and Oscillospiraceae, were significantly less abundant while Verrucomicrobiota, particularly the family Akkermansiaceae, were increasingly more prevalent during peak infection in mice challenged with a high virus dose. In contrast to the cecal microbiome, the lung microbiome showed similar microbial diversity among the control, low and high challenge virus groups, independent of antiviral treatment. Bacterial phyla in the lungs such as Bacteroidota decreased while Firmicutes and Proteobacteria were significantly enriched in mice challenged with a high dose of SARS-CoV-2. In summary, we identified changes in the cecal and lung microbiome of K18-hACE2 mice with severe clinical signs of SARS-CoV-2 infection. IMPORTANCEThe COVID-19 pandemic has resulted in millions of deaths. The hosts respiratory and intestinal microbiome can affect directly or indirectly the immune system during viral infections. We characterized the cecal and lung microbiome in a relevant mouse model challenged with a low and high dose of SARS-CoV-2 in the presence or absence of an antiviral Mpro inhibitor, GC376. Decreased microbial diversity and taxonomic abundances of the phyla Firmicutes, particularly Lachnospiraceae, correlating with infection dosage was observed in the cecum. In addition, microbes within the family Akkermansiaceae were increasingly more prevalent during peak infection, which is observed in other viral infections. The lung microbiome showed similar microbial diversity to the control, independent of antiviral treatment. Decreased Bacteroidota and increased Firmicutes and Proteobacteria were observed in the lungs in a virus dose-dependent manner. These studies add to a better understanding of the complexities associated with the intestinal microbiome during respiratory infections.

microbiology↗