Protective efficacy of recombinant turkey herpes virus (rHVT-H5) and inactivated H5N1 vaccines in commercial Mulard ducks against the highly pathogenic avian influenza (HPAI) H5N1 clade 2.2.1 virus

In Egypt, ducks kept for commercial purposes constitute the second highest poultry population, at 150 million ducks/year. Hence, ducks play an important role in the introduction and transmission of avian influenza (AI) in the Egyptian poultry population. Attempts to control outbreaks include the use of vaccines, which have varying levels of efficacy and failure. To date, the effects of vaccine efficacy has rarely been determined in ducks. In this study, we evaluated the protective efficacy of a live recombinant vector vaccine based on a turkey Herpes Virus (HVT) expressing the H5 gene from a clade 2.2 H5N1 HPAIV strain (A/Swan/ Hungary/499/2006) (rHVT-H5) and a bivalent inactivated H5N1 vaccine prepared from clade 2.2.1 and 2.2.1.1 H5N1 seeds in Mulard ducks. A 0.3ml/dose subcutaneous injection of rHVT-H5 vaccine was administered to one-day-old ducklings (D1) and another 0.5ml/ dose subcutaneous injection of the inactivated MEFLUVAC was administered at 7 days (D7). Four separate challenge experiments were conducted at Days 21, 28, 35 and 42, in which all the vaccinated ducks were challenged with 106EID50/duck of H5N1 HPAI virus (A/ chicken/Egypt/128s/2012(H5N1) (clade 2.2.1) via intranasal inoculation. Maternal-derived antibody regression and post-vaccination antibody immune responses were monitored weekly. Ducks vaccinated at 21, 28, 35 and 42 days with the rHVT-H5 and MEFLUVAC vaccines were protected against mortality (80%, 80%, 90% and 90%) and (50%, 70%, 80% and 90%) respectively, against challenges with the H5N1 HPAI virus. The amount of viral shedding and shedding rates were lower in the rHVT-H5 vaccine groups than in the MEFLUVAC groups only in the first two challenge experiments. However, the non-vaccinated groups shed significantly more of the virus than the vaccinated groups. Both rHVT-H5 and MEFLUVAC provide early protection, and rHVT-H5 vaccine in particular provides protection against HPAI challenge.S1 Table. Weekly mean HI titres (log2 ± SD) using A/Swan/Hungary/4999/2006) rHVT/Ag that indicate the immune response to the rHVT-H5 vaccination. S1 Table legend: Different upper case letters in a row denote the presence of statistically significant (p 0.05) differences. Group I (vaccinated with rHVT-H5 vaccine at 1 day old), Group II (vaccinated with inactivated KV-H5 vaccine at 8 days old), Group III (unvaccinated control).S2 Table. Weekly mean HI titres (log2 ± SD) measured using (A/chicken/Egypt/Q1995D/ 2010) V/H5N1/Ag that indicates the immune response to the KV-H5 vaccination. S2 Table legend: Different upper case letters in a row denote the presence of statistically significant (p 0.05) differences. Group 1 (vaccinated with rHVT-H5 vaccine at 1 day old), Group II (vaccinated with inactivated KV-H5 vaccine at 8 days old), Group III (unvaccinated control).S3 Table. Weekly mean HI titres (log2 ± SD) measured using (A/chicken/Egypt/128S/2012) C/H5N1/Ag that indicates the immune response to the challenge virus. S3 Table legend: Different upper case letters in a row denote the presence of statistically significant (p 0.05) differences. Group 1 (vaccinated with rHVT-H5 vaccine at 1 day old), Group II (vaccinated with inactivated KV-H5 vaccine at 8 days old), Group III (unvaccinated control).This work was supported by the United States Agency for International Development (USAID) under a grant (AID-263-IO-11-00001, Mod. #3) and within the framework of OSRO/EGY/101/ USA, which applies to projects jointly implemented by the FAO, GOVS and NLQP.http://www.plosone.orgam2016Production Animal StudiesVeterinary Tropical Disease

Protective efficacy of recombinant turkey herpes virus (rHVT-H5) and inactivated H5N1 vaccines in commercial Mulard ducks against the highly pathogenic avian influenza (HPAI) H5N1 clade 2.2.1 virus

In Egypt, ducks kept for commercial purposes constitute the second highest poultry population, at 150 million ducks/year. Hence, ducks play an important role in the introduction and transmission of avian influenza (AI) in the Egyptian poultry population. Attempts to control outbreaks include the use of vaccines, which have varying levels of efficacy and failure. To date, the effects of vaccine efficacy has rarely been determined in ducks. In this study, we evaluated the protective efficacy of a live recombinant vector vaccine based on a turkey Herpes Virus (HVT) expressing the H5 gene from a clade 2.2 H5N1 HPAIV strain (A/Swan/ Hungary/499/2006) (rHVT-H5) and a bivalent inactivated H5N1 vaccine prepared from clade 2.2.1 and 2.2.1.1 H5N1 seeds in Mulard ducks. A 0.3ml/dose subcutaneous injection of rHVT-H5 vaccine was administered to one-day-old ducklings (D1) and another 0.5ml/ dose subcutaneous injection of the inactivated MEFLUVAC was administered at 7 days (D7). Four separate challenge experiments were conducted at Days 21, 28, 35 and 42, in which all the vaccinated ducks were challenged with 106EID50/duck of H5N1 HPAI virus (A/ chicken/Egypt/128s/2012(H5N1) (clade 2.2.1) via intranasal inoculation. Maternal-derived antibody regression and post-vaccination antibody immune responses were monitored weekly. Ducks vaccinated at 21, 28, 35 and 42 days with the rHVT-H5 and MEFLUVAC vaccines were protected against mortality (80%, 80%, 90% and 90%) and (50%, 70%, 80% and 90%) respectively, against challenges with the H5N1 HPAI virus. The amount of viral shedding and shedding rates were lower in the rHVT-H5 vaccine groups than in the MEFLUVAC groups only in the first two challenge experiments. However, the non-vaccinated groups shed significantly more of the virus than the vaccinated groups. Both rHVT-H5 and MEFLUVAC provide early protection, and rHVT-H5 vaccine in particular provides protection against HPAI challenge.S1 Table. Weekly mean HI titres (log2 ± SD) using A/Swan/Hungary/4999/2006) rHVT/Ag that indicate the immune response to the rHVT-H5 vaccination. S1 Table legend: Different upper case letters in a row denote the presence of statistically significant (p 0.05) differences. Group I (vaccinated with rHVT-H5 vaccine at 1 day old), Group II (vaccinated with inactivated KV-H5 vaccine at 8 days old), Group III (unvaccinated control).S2 Table. Weekly mean HI titres (log2 ± SD) measured using (A/chicken/Egypt/Q1995D/ 2010) V/H5N1/Ag that indicates the immune response to the KV-H5 vaccination. S2 Table legend: Different upper case letters in a row denote the presence of statistically significant (p 0.05) differences. Group 1 (vaccinated with rHVT-H5 vaccine at 1 day old), Group II (vaccinated with inactivated KV-H5 vaccine at 8 days old), Group III (unvaccinated control).S3 Table. Weekly mean HI titres (log2 ± SD) measured using (A/chicken/Egypt/128S/2012) C/H5N1/Ag that indicates the immune response to the challenge virus. S3 Table legend: Different upper case letters in a row denote the presence of statistically significant (p 0.05) differences. Group 1 (vaccinated with rHVT-H5 vaccine at 1 day old), Group II (vaccinated with inactivated KV-H5 vaccine at 8 days old), Group III (unvaccinated control).This work was supported by the United States Agency for International Development (USAID) under a grant (AID-263-IO-11-00001, Mod. #3) and within the framework of OSRO/EGY/101/ USA, which applies to projects jointly implemented by the FAO, GOVS and NLQP.http://www.plosone.orgam2016Production Animal StudiesVeterinary Tropical Disease

Influenza at the animal-human interface: A review of the literature for virological evidence of human infection with swine or avian influenza viruses other than A(H5N1)

Factors that trigger human infection with animal influenza virus progressing into a pandemic are poorly understood. Within a project developing an evidence-based risk assessment framework for influenza viruses in animals, we conducted a review of the literature for evidence of human infection with animal influenza viruses by diagnostic methods used. The review covering Medline, Embase, SciSearch and CabAbstracts yielded 6,955 articles, of which we retained 89; for influenza A(H5N1) and A(H7N9), the official case counts of the World Health Organization were used. An additional 30 studies were included by scanning the reference lists. Here, we present the findings for confirmed infections with virological evidence. We found reports of 1,419 naturally infected human cases, of which 648 were associated with avian influenza virus (AIV) A(H5N1), 375 with other AIV subtypes, and 396 with swine influenza virus (SIV). Human cases naturally infected with AIV spanned haemagglutinin subtypes H5, H6, H7, H9 and H10. SIV cases were associated with endemic SIV of H1 and H3 subtype d

Highly Pathogenic Avian Influenza Virus Subtype H5N1 in Africa: A Comprehensive Phylogenetic Analysis and Molecular Characterization of Isolates

Highly pathogenic avian influenza virus A/H5N1 was first officially reported in Africa in early 2006. Since the first outbreak in Nigeria, this virus spread rapidly to other African countries. From its emergence to early 2008, 11 African countries experienced A/H5N1 outbreaks in poultry and human cases were also reported in three of these countries. At present, little is known of the epidemiology and molecular evolution of A/H5N1 viruses in Africa. We have generated 494 full gene sequences from 67 African isolates and applied molecular analysis tools to a total of 1,152 A/H5N1 sequences obtained from viruses isolated in Africa, Europe and the Middle East between 2006 and early 2008. Detailed phylogenetic analyses of the 8 gene viral segments confirmed that 3 distinct sublineages were introduced, which have persisted and spread across the continent over this 2-year period. Additionally, our molecular epidemiological studies highlighted the association between genetic clustering and area of origin in a majority of cases. Molecular signatures unique to strains isolated in selected areas also gave us a clearer picture of the spread of A/H5N1 viruses across the continent. Mutations described as typical of human influenza viruses in the genes coding for internal proteins or associated with host adaptation and increased resistance to antiviral drugs have also been detected in the genes coding for transmembrane proteins. These findings raise concern for the possible human health risk presented by viruses with these genetic properties and highlight the need for increased efforts to monitor the evolution of A/H5N1 viruses across the African continent. They further stress how imperative it is to implement sustainable control strategies to improve animal and public health at a global level

Predominance and geo-mapping of avian influenza H5N1 in poultry sectors in Egypt

Highly pathogenic avian influenza (HPAI) virus of the H5N1 subtype has been enzootic in the Egyptian poultry with significant human infections since 2008. This work evaluates the epidemiological and virological information from February 2006 to May 2015 in spatial and temporal terms. Only data with confirmed HPAI H5N1 sub-type were collected, and matched with the epidemiological data from various spatially and temporally-dispersed surveillances implemented between 2006 and 2015. Spatio-temporal analysis was conducted on a total of 3338 confirmed H5N1 HPAI poultry disease outbreaks and outputs described based on transmission patterns, poultry species, production types affected, trade, geographic and temporal distributions in Egypt. The H5N1 virus persists in the Egyptian poultry displaying a seasonal pattern with peak prevalence between January and March. There was no specific geographic pattern, but chickens and ducks were more affected. However, relatively higher disease incidences were recorded in the Nile Delta. Phylogenetic studies of the haemagglutinin gene sequences of H5N1 viruses indicated that multiple clusters circulated between 2006 and 2015, with significant deviations in circulation. Epidemiological dynamics of HPAI has changed with the origins of majority of outbreaks shifted to household poultry. The persistence of HPAI H5N1 in poultry with recurrent and sporadic infections in humans can influence virus evolution spatio-temporally. Household poultry plays significant roles in the H5N1 virus transmission to poultry and humans, but the role of commercial poultry needs further clarifications. While poultry trading supports the persistence and transmission of H5N1, the role of individual species may warrant further investigation. Surveillance activities, applying a multi-sectoral approach, are recommended

Model of experimental design.

<p>Model of experimental design.</p

HI titres^* (mean ±SD) and sero-positivity rates for the selected ducks subject to the four challenge experiments^**.

<p>HI titres^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0156747#t002fn002" target="_blank">*</a>} (mean ±SD) and sero-positivity rates for the selected ducks subject to the four challenge experiments^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0156747#t002fn003" target="_blank">**</a>}.</p

Morbidity and mortality parameters measured in animals subjected to four separate challenge experiments (Exp.1, 2, 3 and 4)^*.

<p>Morbidity and mortality parameters measured in animals subjected to four separate challenge experiments (Exp.1, 2, 3 and 4)^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0156747#t003fn001" target="_blank">*</a>}.</p

Weekly HI mean titres (log2 ± SD) using (A/chicken/Mexico/232/1994) H5N2/Ag that indicate the MDAs’ regression profile.

<p>Weekly HI mean titres (log2 ± SD) using (A/chicken/Mexico/232/1994) H5N2/Ag that indicate the MDAs’ regression profile.</p

Weekly mean HI titres (log2 ± SD) showing the results of MDA regression profiles and post vaccination immune responses in different experimental groups*.

<p>Weekly Mean HI titers (log 2) measured using (<b>1a</b>) (A/Chicken/Mexico/232/1994) H5N2/Ag indicating MDA regression; (<b>1b</b>) (A/Swan/Hungary/4999/2006)rHVT-H5/Ag indicating immune response to rHVT-H5 vaccination; (<b>1c</b>) (A/Chicken/Egypt/Q1995D/2010)H5N1/Ag indicating immune response to KVT-H5 vaccination; (<b>1d</b>) (A/H5N1/Chicken/Egypt/128s/2012)C/H5N1/Ag indicating immune response to challenge virus antigen. *<i>Group I (1b) (vaccinated with rHVT-H5 vaccine at 1 day old)</i>, <i>Group II (1c) (vaccinated with inactivated MEFLUVAC-H5 vaccine at 8 days old)</i>, <i>Group III (1d) (unvaccinated control)</i>.</p