Impact of sex on humoral immunity with live influenza B virus vaccines in mice.

Influenza B virus (FLUBV) poses a significant infectious threat, with frequent vaccine mismatch limiting its effectiveness. Our previous work investigated the safety and efficacy of modified live attenuated FLUBV vaccines with rearranged genomes (FluB-RAM and FluB-RANS) or a temperature-sensitive PB1 segment with a C-terminal HA tag (FluB-att). In this study, we compared the immune responses of female and male DBA/2J mice vaccinated with these vaccines, including versions containing a chimeric HA segment with an N-terminal IgA-inducing peptide (IGIP). Importantly, both recombinant viruses with and without IGIP remained genetically stable during egg passage. We found that introducing IGIP strengthened vaccine attenuation, particularly for FluB-RAM/IGIP. Prime-boost vaccination completely protected mice against lethal challenge with a homologous FLUBV strain. Notably, recombinant viruses induced robust neutralizing antibody responses (hemagglutination inhibition titers ≥40) alongside antibodies against NA and NP. Interestingly, female mice displayed a consistent trend of enhanced humoral and cross-reactive IgG and IgA responses against HA, NA, and NP compared to male counterparts, regardless of the vaccine used. However, the presence of IGIP generally led to lower anti-HA responses but higher anti-NA and anti-NP responses, particularly of the IgA isotype. These trends were further reflected in mucosal and serological responses two weeks after challenge, with clear distinctions based on sex, vaccine backbone, and IGIP inclusion. These findings hold significant promise for advancing the development of universal influenza vaccines.

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 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.

Influenza antivirals and animal models.

Influenza A and B viruses are among the most prominent human respiratory pathogens. About 3-5 million severe cases of influenza are associated with 300 000-650 000 deaths per year globally. Antivirals effective at reducing morbidity and mortality are part of the first line of defense against influenza. FDA-approved antiviral drugs currently include adamantanes (rimantadine and amantadine), neuraminidase inhibitors (NAI; peramivir, zanamivir, and oseltamivir), and the PA endonuclease inhibitor (baloxavir). Mutations associated with antiviral resistance are common and highlight the need for further improvement and development of novel anti-influenza drugs. A summary is provided for the current knowledge of the approved influenza antivirals and antivirals strategies under evaluation in clinical trials. Preclinical evaluations of novel compounds effective against influenza in different animal models are also discussed.

Reverse Genetics for Influenza A and B Viruses Driven by Swine Polymerase I Promoter.

Influenza viruses are considered prominent pathogens of humans and animals that are extensively investigated because of public health importance. Plasmid-based reverse genetics is a fundamental tool that facilitates the generation of genetically modified viruses from a cDNA copy. The ability to rescue viruses enables researchers to understand different biological characteristics including IV replication, pathogenesis, and transmission. Furthermore, understanding the biology and ability to manipulate different aspects of the virus can aid in providing a better understanding of the mechanisms of antiviral resistance and development of alternative vaccination strategies. This chapter describes the process of cloning cDNA copies of IAV and IBV RNA segments into a swine polymerase-driven reverse genetics plasmid vector, successful generation of recombinant IVs in swine cells, and propagation of virus in cells or eggs. The swine polymerase reverse genetics system was previously shown to be efficient for de novo rescue of human-, swine-, and avian-origin IAVs and IBV in swine and human origin cell lines utilizing the same protocols discussed in this chapter.

A Novel Recombinant Newcastle Disease Vaccine Improves Post- In Ovo Vaccination Survival with Sustained Protection against Virulent Challenge.

In ovo vaccination has been employed by the poultry industry for over 20 years to control numerous avian diseases. Unfortunately, in ovo live vaccines against Newcastle disease have significant limitations, including high embryo mortality and the inability to induce full protection during the first two weeks of life. In this study, a recombinant live attenuated Newcastle disease virus vaccine containing the antisense sequence of chicken interleukin 4 (IL-4), rZJ1*L-IL4R, was used. The rZJ1*L-IL4R vaccine was administered in ovo to naïve specific pathogen free embryonated chicken eggs (ECEs) and evaluated against a homologous challenge. Controls included a live attenuated recombinant genotype VII vaccine based on the virus ZJ1 (rZJ1*L) backbone, the LaSota vaccine and diluent alone. In the first of two experiments, ECEs were vaccinated at 18 days of embryonation (DOE) with either 104.5 or 103.5 50% embryo infectious dose (EID50/egg) and chickens were challenged at 21 days post-hatch (DPH). In the second experiment, 103.5 EID50/egg of each vaccine was administered at 19 DOE, and chickens were challenged at 14 DPH. Chickens vaccinated with 103.5 EID50/egg of rZJ1*L-IL4R had hatch rates comparable to the group that received diluent alone, whereas other groups had significantly lower hatch rates. All vaccinated chickens survived challenge without displaying clinical disease, had protective hemagglutination inhibition titers, and shed comparable levels of challenge virus. The recombinant rZJ1*L-IL4R vaccine yielded lower post-vaccination mortality rates compared with the other in ovo NDV live vaccine candidates as well as provided strong protection post-challenge.

Rational design of a deuterium-containing M2-S31N channel blocker UAWJ280 with in vivo antiviral efficacy against both oseltamivir sensitive and -resistant influenza A viruses.

Seasonal influenza A virus (IAV) infections are among the most important global health problems. FDA-approved antiviral therapies against IAV include neuraminidase inhibitors, M2 inhibitors, and polymerase inhibitor baloxavir. Resistance against adamantanes (amantadine and rimantadine) is widespread as virtually all IAV strains currently circulating in the human population are resistant to adamantanes through the acquisition of the S31N mutation. The neuraminidase inhibitor-resistant strains also contain the M2-S31N mutant, suggesting M2-S31N is a high-profile antiviral drug target. Here we report the development of a novel deuterium-containing M2-S31N inhibitor UAWJ280. UAWJ280 had broad-spectrum antiviral activity against both oseltamivir sensitive and -resistant influenza A strains and had a synergistic antiviral effect in combination with oseltamivir in cell culture. In vivo pharmacokinetic (PK) studies demonstrated that UAWJ280 had favourable PK properties. The in vivo mouse model study showed that UAWJ280 was effective alone or in combination with oseltamivir in improving clinical signs and survival after lethal challenge with an oseltamivir sensitive IAV H1N1 strain. Furthermore, UAWJ280 was also able to ameliorate clinical signs and increase survival when mice were challenged with an oseltamivir-resistant IAV H1N1 strain. In conclusion, we show for the first time that the M2-S31N channel blocker UAWJ280 has in vivo antiviral efficacy in mice that are infected with either oseltamivir sensitive or -resistant IAVs, and it has a synergistic antiviral effect with oseltamivir.

Mutation E48K in PB1 Polymerase Subunit Improves Stability of a Candidate Live Attenuated Influenza B Virus Vaccine.

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.

FluB-RAM and FluB-RANS: Genome Rearrangement as Safe and Efficacious Live Attenuated Influenza B Virus Vaccines.

Influenza B virus (IBV) is considered a major respiratory pathogen responsible for seasonal respiratory disease in humans, particularly severe in children and the elderly. Seasonal influenza vaccination is considered the most efficient strategy to prevent and control IBV infections. Live attenuated influenza virus vaccines (LAIVs) are thought to induce both humoral and cellular immune responses by mimicking a natural infection, but their effectiveness has recently come into question. Thus, the opportunity exists to find alternative approaches to improve overall influenza vaccine effectiveness. Two alternative IBV backbones were developed with rearranged genomes, rearranged M (FluB-RAM) and a rearranged NS (FluB-RANS). Both rearranged viruses showed temperature sensitivity in vitro compared with the WT type B/Bris strain, were genetically stable over multiple passages in embryonated chicken eggs and were attenuated in vivo in mice. In a prime-boost regime in naïve mice, both rearranged viruses induced antibodies against HA with hemagglutination inhibition titers considered of protective value. In addition, antibodies against NA and NP were readily detected with potential protective value. Upon lethal IBV challenge, mice previously vaccinated with either FluB-RAM or FluB-RANS were completely protected against clinical disease and mortality. In conclusion, genome re-arrangement renders efficacious LAIV candidates to protect mice against IBV.

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 microbiomes of SARS-CoV-2-challenged K18-hACE2 transgenic mice in the presence or absence of treatment with the Mpro inhibitor GC-376. Cecum microbiome showed decreased Shannon and inverse (Inv) Simpson diversity indexes 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 Verrucomicrobia, 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-dose challenge virus groups, independent of antiviral treatment. Bacterial phyla in the lungs such as Bacteroidetes 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 microbiomes of K18-hACE2 mice with severe clinical signs of SARS-CoV-2 infection. IMPORTANCE The COVID-19 pandemic has resulted in millions of deaths. The host's respiratory and intestinal microbiome can affect directly or indirectly the immune system during viral infections. We characterized the cecal and lung microbiomes in a relevant mouse model challenged with a low or high dose of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the presence or absence of an antiviral Mpro inhibitor, GC-376. Decreased microbial diversity and taxonomic abundances of the phyla Firmicutes, particularly, Lachnospiraceae, correlating with infection dosage were 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 that of the control, independent of antiviral treatment. Decreased Bacteroidetes 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.

A New Master Donor Virus for the Development of Live-Attenuated Influenza B Virus Vaccines.

Influenza B viruses (IBV) circulate annually, with young children, the elderly and immunocompromised individuals being at high risk. Yearly vaccinations are recommended to protect against seasonally influenza viruses, including IBV. Live attenuated influenza vaccines (LAIV) provide the unique opportunity for direct exposure to the antigenically variable surface glycoproteins as well as the more conserved internal components. Ideally, LAIV Master Donor Viruses (MDV) should accurately reflect seasonal influenza strains. Unfortunately, the continuous evolution of IBV have led to significant changes in conserved epitopes compared to the IBV MDV based on B/Ann Arbor/1/1966 strain. Here, we propose a recent influenza B/Brisbane/60/2008 as an efficacious MDV alternative, as its internal viral proteins more accurately reflect those of circulating IBV strains. We introduced the mutations responsible for the temperature sensitive (ts), cold adapted (ca) and attenuated (att) phenotype of B/Ann Arbor/1/1966 MDV LAIV into B/Brisbane/60/2008 to generate a new MDV LAIV. In vitro and in vivo analysis demonstrated that the mutations responsible of the ts, ca, and att phenotype of B/Ann Arbor/1/1966 MDV LAIV were able to infer the same phenotype to B/Brisbane/60/2008, demonstrating its potential as a new MDV for the development of LAIV to protect against contemporary IBV strains.

Development of a Novel Live Attenuated Influenza A Virus Vaccine Encoding the IgA-Inducing Protein.

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.

Efficacy of GC-376 against SARS-CoV-2 virus infection in the K18 hACE2 transgenic mouse model.

The COVID-19 pandemic caused by the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is the defining global health emergency of this century. GC-376 is a Mpro inhibitor with antiviral activity against SARS-CoV-2 in vitro. Using the K18-hACE2 mouse model, the in vivo antiviral efficacy of GC-376 against SARS-CoV-2 was evaluated. GC-376 treatment was not toxic in K18-hACE2 mice. Overall outcome of clinical symptoms and survival upon SARS-CoV-2 challenge were not improved in mice treated with GC-376 compared to controls. The treatment with GC-376 slightly improved survival from 0 to 20% in mice challenged with a high virus dose at 105 TCID50/mouse. Most notably, GC-376 treatment led to milder tissue lesions, reduced viral loads, fewer presence of viral antigen, and reduced inflammation in comparison to vehicle-treated controls in mice challenged with a low virus dose at 103 TCID50/mouse. This was particularly the case in the brain where a 5-log reduction in viral titers was observed in GC-376 treated mice compared to vehicle controls. This study supports the notion that GC-376 represents a promising lead candidate for further development to treat SARS-CoV-2 infection and that the K18-hACE2 mouse model is suitable to study antiviral therapies against SARS-CoV-2.

Efficacy of GC-376 against SARS-CoV-2 virus infection in the K18 hACE2 transgenic mouse model.

The COVID-19 pandemic caused by the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is the defining global health emergency of this century. GC-376 is a M pro inhibitor with antiviral activity against SARS-CoV-2 in vitro . Using the K18-hACE2 mouse model, the in vivo antiviral efficacy of GC-376 against SARS-CoV-2 was evaluated. GC-376 treatment was not toxic in K18-hACE2 mice and produced milder tissue lesions, reduced viral loads, fewer presence of viral antigen, and reduced inflammation in comparison to vehicle-treated controls, most notably in the brain in mice challenged with a low virus dose. Although GC-376 was not sufficient to improve neither clinical symptoms nor survival, it did show a positive effect against SARS-CoV-2 in vivo . This study supports the notion that the K18-hACE2 mouse model is suitable to study antiviral therapies against SARS-CoV-2, and GC-376 represents a promising lead candidate for further development to treat SARS-CoV-2 infection.

Development of a swine RNA polymerase I driven Influenza reverse genetics system for the rescue of type A and B Influenza viruses.

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.

Reverse genetics for influenza B viruses and recent advances in vaccine development.

Influenza B virus is a respiratory pathogen that affects more severely the pediatric and elderly populations. There are two lineages of influenza B virus that seem to have differential predilection for age groups. Both lineages can co-circulate during the influenza season however one is usually more prominent than the other depending on the season. There are no defined indicators to predict which lineage will dominate in any given season. In recent years, the addition of viruses from both lineages to the seasonal influenza vaccine formulation has improved vaccine protection, although quadrivalent vaccines are not available worldwide. Reverse genetics has facilitated advancements in the field of vaccine development against influenza B virus. Different strategies have been explored showing promising results that could potentially lead to the development broadly protective influenza B virus vaccines.

Maternally-Derived Antibodies Protect against Challenge with Highly Pathogenic Avian Influenza Virus of the H7N3 Subtype.

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 (-)). Both MDA (+) and MDA (-) 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 (-) 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.

Tropism of Newcastle disease virus strains for chicken neurons, astrocytes, oligodendrocytes, and microglia.

BACKGROUND: Newcastle disease (ND), which is caused by infections of poultry species with virulent strains of Avian orthoavulavirus-1, also known as avian paramyxovirus 1 (APMV-1), and formerly known as Newcastle disease virus (NDV), may cause neurological signs and encephalitis. Neurological signs are often the only clinical signs observed in birds infected with neurotropic strains of NDV. Experimental infections have shown that the replication of virulent NDV (vNDV) strains is in the brain parenchyma and is possibly confined to neurons and ependymal cells. However, little information is available on the ability of vNDV strains to infect subset of glial cells (astrocytes, oligodendrocytes, and microglia). The objective of this study was to evaluate the ability of NDV strains of different levels of virulence to infect a subset of glial cells both in vitro and in vivo. Thus, neurons, astrocytes and oligodendrocytes from the brains of day-old White Leghorn chickens were harvested, cultured, and infected with both non-virulent (LaSota) and virulent, neurotropic (TxGB) NDV strains. To confirm these findings in vivo, the tropism of three vNDV strains with varying pathotypes (SA60 [viscerotropic], TxGB [neurotropic], and Tx450 [mesogenic]) was assessed in archived formalin-fixed material from day-old chicks inoculated intracerebrally. RESULTS: Double immunofluorescence for NDV nucleoprotein and cellular markers showed that both strains infected at least 20% of each of the cell types (neurons, astrocytes, and oligodendrocytes). At 24 h post-inoculation, TxGB replicated significantly more than LaSota. Double immunofluorescence (DIFA) with markers for neurons, astrocytes, microglia, and NDV nucleoprotein detected the three strains in all three cell types at similar levels. CONCLUSION: These data indicate that similar to other paramyxoviruses, neurons and glial cells (astrocytes, oligodendrocytes, and microglia) are susceptible to vNDV infection, and suggest that factors other than cellular tropism are likely the major determinant of the neurotropic phenotype.

Alternative Strategy for a Quadrivalent Live Attenuated Influenza Virus Vaccine.

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.

Draft Genome Sequences of Three Ochrobactrum spp. Isolated from Different Avian Hosts in Pakistan.

Here, we present the draft genome sequences of three Ochrobactrum sp. strains with multidrug-resistant properties, isolated in 2015 from a pigeon and two chickens in Pakistan.

Draft Genome Sequences of Five Novel Ochrobactrum spp. Isolated from Different Avian Hosts in Nigeria.

Here, we present the draft genome sequences of five multidrug-resistant novel Ochrobactrum species strains isolated from a pigeon, a duck, and chickens from Nigeria in 2009.