A #nanoemulsion-adjuvanted intranasal #H5N1 #influenza #vaccine protects ferrets against homologous and heterologous H5N1 lethal challenge (Vaccine, abstract)

[Source: US National Library of Medicine, full page: (LINK). Abstract, edited.]

Vaccine. 2019 Sep 5. pii: S0264-410X(19)31147-8. doi: 10.1016/j.vaccine.2019.08.071. [Epub ahead of print]

A nanoemulsion-adjuvanted intranasal H5N1 influenza vaccine protects ferrets against homologous and heterologous H5N1 lethal challenge.

Smith D1, Streatfield SJ2, Acosta H1, Ganesan S1, Fattom A3.

Author information: 1 BlueWillow Biologics, Ann Arbor, MI, United States. 2 Fraunhofer USA Center for Molecular Biotechnology (FhCMB), Newark, DE, United States. 3 BlueWillow Biologics, Ann Arbor, MI, United States. Electronic address: ali.fattom@bluewillow.com.




Flu vaccines administered intramuscularly (IM) have shown seasonally fluctuating efficacy, 20-60%, throughout the last 15 years. We formulated a recombinant H5 (rH5) in our Nanovax® (NE01) (rH5/NE01) adjuvant for intranasal vaccination in ferrets. We evaluated the regimen, one vs two immunization, and cross clade protection a ferret challenge model.


Plant derived recombinant H5 (rH5) antigen was formulated with NE01 and administered intranasally to ferrets. Immunogenicity (IgG), hemagglutination inhibition (HI), and protection against lethal challenge, were measured following one or two immunizations. Protection against homologous (strain A/Indo) and heterologous (strain A/Vn) was evaluated in ferrets following two immunizations.


IN immunization with rH5/NE01 induced significant IgG levels after one and two immunizations. One vaccination did not induce any HI while low HI was measured after two immunizations. Homologous challenge with H5N1 A/ Indonesia showed 100% survival, with minimal weight loss in animals vaccinated twice compared to the unvaccinated controls. Analysis of nasal wash from these challenged ferrets vaccinated twice showed decreased viral shedding compared to unvaccinated controls. Interestingly, animals that received one vaccination showed 88% survival with moderate weight loss. Cross clade protection was evaluated using an increased antigen dose (45 µg rH5). Vaccinated animals demonstrated increased IgG and HAI antibody responses. Both homologous (A/Indo) and heterologous challenge (A/Vietnam) following two immunizations showed 100% survival with no loss of body weight. However viral clearance was more rapid against the homologous (day 3) compared to the heterologous (day 5) post challenge.


Intranasal administration of NE01 adjuvant-formulated rH5 vaccine elicited systemic and probably mucosal immunity that conferred protection against lethal challenge with homologous or heterologous viral strains. It also enhanced viral clearance with decreased shedding. These outcomes strongly suggest that intranasal immunization using NE01 against flu infections warrants clinical testing.

Copyright © 2019. Published by Elsevier Ltd.

KEYWORDS: Ferrets model; Intranasal immunization; Nanoemulsion; Pandemic flu; Vaccine

PMID: 31495593 DOI: 10.1016/j.vaccine.2019.08.071

Keywords: Avian Influenza; H5N1; Vaccines; Animal models.



#Fireworks-like #Surveillance #Approach: The case of HPAI #H5N1 in #WildBirds in #Europe (Transbound Emerg Dis., abstract)

[Source: US National Library of Medicine, full page: (LINK). Abstract, edited.]

Transbound Emerg Dis. 2019 Sep 3. doi: 10.1111/tbed.13342. [Epub ahead of print]

Fireworks-like Surveillance Approach: The case of HPAI H5N1 in Wild Birds in Europe.

Pereira H1,2,3, Artois M1, Bicout DJ1,4.

Author information: 1 VetAgro Sup, Veterinary Campus of Lyon, 69280, Marcy l’Étoile, France. 2 Assistance Publique-Hôpitaux de Paris, Hôpital Européen Georges-Pompidou, Unité de Recherche Clinique, Paris, France. 3 INSERM, Centre d’Investigation Clinique 1418 (CIC1418), Paris, France. 4 Biomathematics and Epidemiology, EPSP- Labo TIMC, UMR 5525, CNRS, Grenoble Alpes University and VetAgro Sup, 69280, Marcy l’Étoile, France.



Highly pathogenic avian influenza (HPAI) risk management requires efficient surveillance of the infection in wild birds for early warning purposes. In this study, our aim was to describe the spread of continent-wide infection cases using a fireworks model and thereby improve current surveillance systems. The fireworks model is a metaphor illustrating the spread of HPAI as a point source epizootic. The approach is based on early detection of the outbreak seeds (sparks from the fireworks) and uses a predictive model of the probability of the occurrence of new cases following a seed introduction; this then determines the spatiotemporal perimeter for intense surveillance investigations. For a case study, we used surveillance data on HPAI H5N1 in wild birds across Europe between 2005 and 2010 to describe the outbreaks and determine the success of the case detection used to inform management of the disease. The fireworks description assumes simultaneous introductions of “seeds” of cases, which then “explode” in local foci but do not merge into a progressive disease wave. This model fit the data well. Using this predictive approach for HPAI cases in EU countries, we found that the investigation radius needed to achieve a detection level of 90% of new cases after an outbreak ranged from 10 km to more than 300 km, depending on the outbreak pattern. Based on these findings, the fireworks approach can be a valuable method for identifying the perimeters and risk areas to be targeted for enhanced surveillance. The rationale of the fireworks approach is quite generic and can easily be adapted to different situations and contexts.

This article is protected by copyright. All rights reserved. © 2019 Blackwell Verlag GmbH.

KEYWORDS: H5N1; Highly pathogenic avian influenza; fireworks-based surveillance; mathematical modelling; wild birds

PMID: 31482660 DOI: 10.1111/tbed.13342

Keywords: Avian Influenza; H5N1; Wild Birds; European Region.


Protective efficacy of an inactivated chimeric #H5 #avian #influenza #vaccine against H5 highly pathogenic avian influenza virus clades and (J Gen Virol., abstract)

[Source: US National Library of Medicine, full page: (LINK). Abstract, edited.]

J Gen Virol. 2018 Dec;99(12):1600-1607. doi: 10.1099/jgv.0.001140. Epub 2018 Oct 25.

Protective efficacy of an inactivated chimeric H5 avian influenza vaccine against H5 highly pathogenic avian influenza virus clades and

Li J1, Hou G1, Wang Y2, Wang S1, Cheng S1, Peng C1, Jiang W1.

Author information: 1 ​China Animal Health and Epidemiology Center, Qingdao, PR China. 2 ​Shanghai Entry-Exit Inspection and Quarantine Bureau, Shanghai, PR China.



The H5 subtype of highly pathogenic avian influenza (HPAI) viruses pose a serious challenge to public health and the poultry industry in China. In this study, we generated a chimeric QH/KJ recombinant virus expressing the entire haemagglutinin (HA)-1 region of the HPAI virus A/chicken/China/QH/2017(H5N6) (clade and the HA2 region of the HPAI virus A/chicken/China/KJ/2017(H5N1) (clade The resulting chimeric PR8-QH/KJ virus exhibited similar in vitro growth kinetics as the parental PR8-QH and PR8-KJ viruses. The chimeric PR8-QH/KJ virus induced specific, cross-reactive haemagglutination-inhibition and serum-neutralizing antibodies against both QH and KJ viruses, although PR8-QH and PR8-KJ exhibited no cross-reactivity with each other. Furthermore, the chimeric PR8-QH/KJ vaccine significantly reduced virus shedding and completely protected chickens from challenge with HPAI H5N6 and H5N1 viruses. However, the Re-8 vaccine against clade viruses provided specific-pathogen-free chickens only partial protection when challenged with QH virus. Our results suggest that the antigenic variation of these epidemic viruses occurred and they can escape the current vaccine immunization. The Re-8 vaccine needs an update. The chimeric PR8-QH/KJ vaccine is effective against H5 HPAI virus clades and in chickens.

KEYWORDS: Avian influenza virus; H5N1; H5N6; chimeric vaccine

PMID: 30358528 DOI: 10.1099/jgv.0.001140 [Indexed for MEDLINE]

Keywords: Avian Influenza; H5N6; H5N1; Poultry; Vaccines.


Structures of #influenza A virus #RNA #polymerase offer insight into viral #genome #replication (Nature, abstract)

[Source: Nature, full page: (LINK). Abstract, edited.]

Structures of influenza A virus RNA polymerase offer insight into viral genome replication

Haitian Fan, Alexander P. Walker, Loïc Carrique, Jeremy R. Keown, Itziar Serna Martin, Dimple Karia, Jane Sharps, Narin Hengrung, Els Pardon, Jan Steyaert, Jonathan M. Grimes & Ervin Fodor

Nature (2019)



Influenza A viruses are responsible for seasonal epidemics, and pandemics can arise from the transmission of novel zoonotic influenza A viruses to humans1,2. Influenza A viruses contain a segmented negative-sense RNA genome, which is transcribed and replicated by the viral-RNA-dependent RNA polymerase (FluPolA) composed of PB1, PB2 and PA subunits3,4,5. Although the high-resolution crystal structure of FluPolA of bat influenza A virus has previously been reported6, there are no complete structures available for human and avian FluPolA. Furthermore, the molecular mechanisms of genomic viral RNA (vRNA) replication—which proceeds through a complementary RNA (cRNA) replicative intermediate, and requires oligomerization of the polymerase7,8,9,10—remain largely unknown. Here, using crystallography and cryo-electron microscopy, we determine the structures of FluPolA from human influenza A/NT/60/1968 (H3N2) and avian influenza A/duck/Fujian/01/2002 (H5N1) viruses at a resolution of 3.0–4.3 Å, in the presence or absence of a cRNA or vRNA template. In solution, FluPolA forms dimers of heterotrimers through the C-terminal domain of the PA subunit, the thumb subdomain of PB1 and the N1 subdomain of PB2. The cryo-electron microscopy structure of monomeric FluPolA bound to the cRNA template reveals a binding site for the 3′ cRNA at the dimer interface. We use a combination of cell-based and in vitro assays to show that the interface of the FluPolA dimer is required for vRNA synthesis during replication of the viral genome. We also show that a nanobody (a single-domain antibody) that interferes with FluPolA dimerization inhibits the synthesis of vRNA and, consequently, inhibits virus replication in infected cells. Our study provides high-resolution structures of medically relevant FluPolA, as well as insights into the replication mechanisms of the viral RNA genome. In addition, our work identifies sites in FluPolA that could be targeted in the development of antiviral drugs.

Keywords: Influenza A; H3N2; H5N1; Viral pathogenesis.


Comparative #pathological #findings in mute #swans (Cygnus olor) naturally infected with highly pathogenic #Avian #influenza viruses #H5N1 and #H5N8 in #Serbia (Vet Ital., abstract)

[Source: US National Library of Medicine, full page: (LINK). Abstract, edited.]

Vet Ital. 2019 Mar 31;55(1):95-101. doi: 10.12834/VetIt.1463.7919.2.

Comparative pathological findings in mute swans (Cygnus olor) naturally infected with highly pathogenic Avian influenza viruses H5N1 and H5N8 in Serbia.

Božić B1, Vučićević I, Polaček V, Vasković N, Petrović T, Pajić M, Aleksić-Kovačević S.

Author information: 1 Scientific Veterinary Institute “Novi Sad”, Rumenački put 20, Novi Sad, Serbia. biljana@niv.ns.ac.rs.



The aim of this study was to compare pathological lesions and viral antigen expression in the organs of mute swans (Cygnus olor) naturally infected with highly pathogenic avian influenza virus subtypes H5N1 and H5N8. The examination was conducted on the carcasses of 22 mute swans which died during the avian influenza outbreaks in Serbia in 2006 and 2016-2017. Avian influenza virus subtype H5N8 isolated from mute swans in 2016-2017 was clustered within the clade group B. After necropsy, lung, liver, spleen, pancreas, kidney and brain tissues were sampled for histopathology and immunohistochemical examination. Avian influenza virus nucleoprotein polyclonal antibodies were used for detecting the viral antigen in the examined tissues. The most significant gross lesions were necrosis and haemorrhages in the pancreas. Major histological lesions were multifocal necroses in the pancreas, spleen and liver, non-purulent encephalitis, lung congestion and oedema. Immunohistochemical demonstration of HPAIV nucleoprotein in pancreas and brain was strongly consistent with histological lesions in both infected groups. Our findings showed that pancreas was the most affected organ in all examined mute swans. In addition to increased mortality rate, similar pathological findings were detected in mute swans naturally infected with highly pathogenic avian influenza viruses H5N1 and H5N8.

PMID: 30951187 DOI: 10.12834/VetIt.1463.7919.2 [Indexed for MEDLINE] Free full text

Keywords: Avian Influenza; H5N1; H5N8; Wild Birds; Serbia.


Efficacy of #clarithromycin against #H5N1 and #H7N9 #avian #influenza a virus #infection in cynomolgus monkeys (Antiviral Res., abstract)

[Source: US National Library of Medicine, full page: (LINK). Abstract, edited.]

Antiviral Res. 2019 Aug 14:104591. doi: 10.1016/j.antiviral.2019.104591. [Epub ahead of print]

Efficacy of clarithromycin against H5N1 and H7N9 avian influenza a virus infection in cynomolgus monkeys.

Arikata M1, Itoh Y2, Shichinohe S3, Nakayama M3, Ishigaki H3, Kinoshita T3, Le MQ4, Kawaoka Y5, Ogasawara K6, Shimizu T1.

Author information: 1 Department of Otorhinolaryngology, Shiga University of Medical Science, Japan. 2 Division of Pathology and Disease Regulation, Department of Pathology, Shiga University of Medical Science, Japan. Electronic address: yasushii@belle.shiga-med.ac.jp. 3 Division of Pathology and Disease Regulation, Department of Pathology, Shiga University of Medical Science, Japan. 4 National Institute of Hygiene and Epidemiology, Hanoi, Viet Nam. 5 Division of Virology, Department of Microbiology and Immunology, Institute of Medical Science, University of Tokyo, Tokyo, Japan; Department of Pathobiological Sciences, University of Wisconsin-Madison, Madison, WI, USA. 6 Division of Pathology and Disease Regulation, Department of Pathology, Shiga University of Medical Science, Japan; Research Center for Animal Life Science, Shiga University of Medical Science, Otsu, Japan.



Clarithromycin (CAM), a 14-membered ring macrolide, has anti-inflammatory and immunomodulatory actions and antiviral effects in seasonal influenza virus infection. We examined the prophylactic and therapeutic efficacy of CAM against H5N1 highly pathogenic and H7N9 low pathogenic avian influenza virus infections in cynomolgus monkeys. CAM suppressed H5N1 virus-induced severe signs of disease in the treated monkeys and inhibited virus propagation in tracheal samples and the production of inflammatory cytokines in the lungs of monkeys infected with H5N1 and H7N9 viruses. The prophylactic administration of CAM showed more suppressive effects on clinical signs of disease and viral titers than did therapeutic administration. Thus, since administration of CAM alone showed a tendency to ameliorate clinical sings and to reduce levels of inflammatory cytokines, the macrolides are expected to have effects in combination with the other antiviral drugs on the prophylactic and treatment of patients with severe avian influenza virus infection, which should be further investigated.

Copyright © 2019. Published by Elsevier B.V.

KEYWORDS: Avian influenza virus; Clarithromycin; Cynomolgus monkey; H5N1; H7N9

PMID: 31421167 DOI: 10.1016/j.antiviral.2019.104591

Keywords: Antivirals; Antibiotics; Clarithromycin; Avian Influenza; H5N1; H7N9; Animal models.


#Evaluation of novel #disposable #bioreactors on #pandemic #influenza virus production [#CVV] (PLoS One, abstract)

[Source: PLoS One, full page: (LINK). Abstract, edited.]


Evaluation of novel disposable bioreactors on pandemic influenza virus production

Chia-Chun Lai, Tsai-Chuan Weng, Yu-Fen Tseng, Jen-Ron Chiang, Min-Shi Lee, Alan Yung-Chih Hu

Published: August 12, 2019 / DOI: https://doi.org/10.1371/journal.pone.0220803



Since 1997, the highly pathogenic influenza H5N1 virus has spread from Hong Kong. According to the WHO bulletin report, the H5N1 virus is a zoonotic disease threat that has infected more than 850 humans, causing over 450 deaths. In addition, an outbreak of another new and highly pathogenic influenza virus (H7N9) occurred in 2013 in China. These highly pathogenic influenza viruses could potentially cause a worldwide pandemic. it is crucial to develop a rapid production platform to meet this surge demand against any possible influenza pandemic. A potential solution for this problem is the use of cell-based bioreactors for rapid vaccine production. These novel bioreactors, used for cell-based vaccine production, possess various advantages. For example, they enable a short production time, allow for the handling highly pathogenic influenza in closed environments, and can be easily scaled up. In this study, two novel disposable cell-based bioreactors, BelloCell and TideCell, were used to produce H5N1 clade II and H7N9 candidate vaccine viruses (CVVs). Madin-Darby canine kidney (MDCK) cells were used for the production of these influenza CVVs. A novel bench-scale bioreactor named BelloCell bioreactor was used in the study. All culturing conditions were tested and scaled to 10 L industrial-scale bioreactor known as TideCell002. The performances of between BelloCell and TideCell were similar in cell growth, the average MDCK cell doubling time was slightly decreased to 25 hours. The systems yielded approximately 39.2 and 18.0 μg/ml of HA protein with the 10-liter TideCell002 from the H5N1 clade II and H7N9 CVVs, respectively. The results of this study not only highlight the overall effectiveness of these bioreactors but also illustrate the potential of maintaining the same outcome when scaled up to industrial production, which has many implications for faster vaccine production. Although additional studies are required for process optimization, the results of this study are promising and show that oscillating bioreactors may be a suitable platform for pandemic influenza virus production.


Citation: Lai C-C, Weng T-C, Tseng Y-F, Chiang J-R, Lee M-S, Hu AY-C (2019) Evaluation of novel disposable bioreactors on pandemic influenza virus production. PLoS ONE 14(8): e0220803. https://doi.org/10.1371/journal.pone.0220803

Editor: Balaji Manicassamy, University of Iowa, UNITED STATES

Received: February 20, 2019; Accepted: July 23, 2019; Published: August 12, 2019

Copyright: © 2019 Lai et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Data Availability: All relevant data are within the manuscript and its Supporting Information files.

Funding: The authors would like to thank Mr. Anthony Chang for English editing, and the US CDC for supplying the egg-derived vaccine strains. The authors would also like to thank the funding support from the Ministry of Science and Technology (R.O.C. 102-2622-B-400-001-CC2; 103-2622-B-400-001-CC2). Mr. Chia-Chun Lai is currently pursuing his Ph.D. studies under the Graduate Program of Biotechnology in Medicine, National Tsing Hua University and the National Health Research Institutes. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Competing interests: The authors have declared that no competing interests exist.

Keywords: Avian Influenza; H5N1; H7N9; Vaccines; Pandemic preparedness.