Printer Friendly

New Reassortant Clade Avian Influenza A(H5N6) Virus in Wild Birds, South Korea, 2017-18.

Clade H5 highly pathogenic avian influenza viruses (HPAIVs) have evolved by reassortment with different neuraminidase (NA) and internal genes of prevailing low pathogenicity avian influenza viruses (LPAIVs) and other HPAIVs to generate new genotypes and further evolved into genetic subgroups A-D since 2014 (1). Among these, subgroups A and B viruses were disseminated over vast geographic regions by migratory wild birds (2,3). Subgroup B influenza A(H5N8) viruses were detected in Qinghai Lake, China, and Uvs-Nuur Lake, Russia, during May-June 2016 (Qinghai/Uvs-like), followed by the identification of reassortant viruses in multiple Eurasian countries (4-6). Recently, subgroup B H5N6 viruses were isolated from birds in Greece during February 2017 and England, Germany, the Netherlands, Japan, and Taiwan during winter 2017-18 (7,8).

During December 2017-January 2018 in South Korea, we isolated 6 H5N6 HPAIVs from 231 fecal samples of wild birds collected from the banks of the Cheongmi-cheon River (37[degrees]06'56.9"N, 127[degrees]25'18.3"E) and 34 from 222 fecal samples collected from the banks of the Gokgyo-cheon River (36[degrees]45'12.3'N, 127[degrees]07'12.7'E) (online Technical Appendix 1, These wild bird habitats are wintering sites of migratory waterfowl, including mallard (Anas platyrhynchos), spot-billed duck (Anas poecilorhyncha), Mandarin duck (Aix galericulata), and common teal (Anas crecca). The Gokgyo-cheon River is a major habitat site for Mandarin ducks, and numerous HPAIVs were detected in fecal samples from Mandarin ducks during 2011, 2015, and 2016 (9). We identified avian influenza virus-positive fecal samples from 38 Mandarin ducks and 2 mallards, based on DNA barcoding technique (10). We performed full-length genome sequencing and comparative phylogenetic analysis on 19 of the 40 isolates (online Technical Appendix 1; online Technical Appendix 2, article/24/10/18-0461-Techapp2.xlsx).

All H5N6 isolates shared high nucleotide sequence identities in all 8 gene segments (99.58%-100%) and were identified as HPAIVs based on the presence of multiple basic amino acids at the HA proteolytic cleavage site (PLREKRRKR/G). Searches of the GISAID (https:// and BLAST (https://blast.ncbi.nlm.nih. gov/Blast.cgi) databases indicated that all 8 genomes had the highest nucleotide identity with A/Great_Blackbacked_Gull/Netherlands/1/2017 (Netherlands/1) clade subgroup B H5N6 strain from December 2017 (99.17%-99.79%), rather than subgroup B H5N6 viruses from Japan and Taiwan collected during December 2017 (97.18%-99.27%).

In phylogenetic analysis, we identified 2 genotypes of subgroup B H5N6 viruses (online Technical Appendix 1 Figures 1, 2): genotypes B.N6.1 and B.N6.2. The genotype B.N6.1 viruses were identified from South Korea, Japan, Taiwan, Greece, and the Netherlands (Netherlands/1 strain), and the genotype B.N6.2 viruses were detected from England, Germany, and the Netherlands. For genotype B.N6.1, all genes except NA clustered with H5N8 HPAIV of previously reported genotypes, H5N8-NL cluster I in the Netherlands (6), Ger-11-16 in Germany (5), and Duck/Poland/82a/16-like in Italy (4). The NA gene clustered with LPAIVs circulating in wild birds in Eurasia and separated into 2 clusters, suggesting the potential for >2 independent reassortment events between H5N8 virus and unidentified wild bird origin N6 segments. Consistent clustering of South Korea isolates with the Netherlands/1 strain in maximum-likelihood (ML) phylogenies for each gene supported by high ML bootstrap values (86-100) suggests their close relationship. The genotype B.N6.2 viruses had different polymerase basic 2 (PB2) and polymerase acidic (PA) genes from genotype B.N6.1. The polymerase basic 2 gene probably originated from other LPAIVs, and a polymerase acidic gene originated from H5N8-NL cluster II genotype (6,7). The phylogenetic network and ML analysis suggest that H5N6 viruses have evolved from subgroup B H5N8 viruses into 3 independent pathways, detected in Greece, Europe/South Korea, and Japan/Taiwan (online Technical Appendix 1, Figures 2,3).

The time of most recent common ancestry (tMRCA) for each gene of genotype B.N6.1 H5N6 viruses isolated during winter 2017-18 in Eurasia, except for the NA gene, ranged from January 2016 to October 2016, suggesting that genotype B.N6.1 viruses diverged during the previous year. The tMRCA of the NA gene was September 2015 (95% highest posterior density August 2014-August 2016). The tMRCA of the NA gene has wide 95% highest posterior density range because only a few recent N6 genes of LPAIVs were available in databases for analysis. The tMRCA for each gene of H5N6 HPAIVs identified in South Korea ranged from July through September 2017, suggesting that ancestors of these viruses emerged among wild birds during or after summer 2017, possibly at the breeding and molting sites in the Palearctic region (Table; online Technical Appendix Figure 4). Detection of H5N6 HPAIV from fecal samples of wild birds in South Korea during the 2017-18 wintering season and our phylogenetic analysis suggest that the viruses had moved through wild birds during the fall migration season.

On the basis of our data and migratory pattern of birds, we estimate that H5N6 viruses possibly descended from H5N8 viruses circulating during 2016-17, reaching breeding regions of wild birds during early 2017, followed by dissemination into Europe and East Asia during the fall migration. Enhanced surveillance in wild birds is needed for early detection of new introductions of HPAIV and to trace the transmission route of HPAIV.

This work was funded by Konkuk University in 2015.

Mr. Kwon is a PhD candidate at Konkuk University, Seoul, South Korea. His primary research interest is the epidemiology of HPAIVs in wild birds. Ms. Jeong is a PhD candidate at Konkuk University, Seoul. Her primary research interest is the epidemiology of viruses in wild birds.


(1.) Sonnberg S, Webby RJ, Webster RG. Natural history of highly pathogenic avian influenza H5N1. Virus Res. 2013;178:63-77.

(2.) Lee DH, Bertran K, Kwon JH, Swayne DE. Evolution, global spread, and pathogenicity of highly pathogenic avian influenza H5Nx clade J Vet Sci. 2017;18(S1):269-80.

(3.) Global Consortium for H5N8 and Related Influenza Viruses. Role for migratory wild birds in the global spread of avian influenza H5N8. Science. 2016;354:213-7. science.aaf8852

(4.) Fusaro A, Monne I, Mulatti P, Zecchin B, Bonfanti L, Ormelli S, et al. Genetic diversity of highly pathogenic avian influenza A(H5N8/H5N5) viruses in Italy, 2016-17. Emerg Infect Dis. 2017;23:1543-7.

(5.) Pohlmann A, Starick E, Grund C, Hoper D, Strebelow G, Globig A, et al. Swarm incursions of reassortants of highly pathogenic avian influenza virus strains H5N8 and H5N5, clade, Germany, winter 2016/17. Sci Rep. 2018;8:15. s41598-017-16936-8

(6.) Beerens N, Heutink R, Bergervoet SA, Harders F, Bossers A, Koch G. Multiple reassorted viruses as cause of highly pathogenic avian influenza A(H5N8) virus epidemic, the Netherlands, 2016. Emerg Infect Dis. 2017;23:1974-81. eid2312.171062

(7.) Beerens N, Koch G, Heutink R, Harders F, Vries DPE, Ho C, et al. Novel highly pathogenic avian influenza A(H5N6) virus in the Netherlands, December 2017. Emerg Infect Dis. 2018; 24:770-3.

(8.) Liu YP, Lee DH, Chen LH, Lin YJ, Li WC, Hu SC, et al. Detection of reassortant H5N6 clade highly pathogenic avian influenza virus in a black-faced spoonbill (Platalea minor) found dead, Taiwan, 2017. Infect Genet Evol. 2018;62:275-8.

(9.) Kwon JH, Lee DH, Swayne DE, Noh JY, Yuk SS, Erdene-Ochir TO, et al. Reassortant clade avian influenza A(H5N6) virus in a wild Mandarin duck, South Korea, 2016. Emerg Infect Dis. 2017;23:822-6. eid2305.161905

(10.) Lee DH, Lee HJ, Lee YJ, Kang HM, Jeong OM, Kim MC, et al. DNA barcoding techniques for avian influenza virus surveillance in migratory bird habitats. J Wildl Dis. 2010;46:649-54. http://dx.doi.Org/10.7589/0090-3558-46.2.649

Address for correspondence: Chang-Seon Song, Avian Disease Laboratory, College of Veterinary Medicine, Konkuk University, 1 Hwayang-dong, Gwangjin-gu, Seoul, 143-701, South Korea; email:

Jung-Hoon Kwon, [1] Sol Jeong, [1] Dong-Hun Lee, David E. Swayne, Yu-jin Kim, Sun-hak Lee, Jin-Yong Noh, Tseren-Ochir Erdene-Ochir, Jei-Hyun Jeong, Chang-Seon Song

Author affiliations: Konkuk University, Seoul, South Korea (J.-H. Kwon, S. Jeong, Y.-J. Kim, S.-H. Lee, J.-Y. Noh, T.-O. Erdene-Ochir, J.-H. Jeong, C.-S. Song); US Department of Agriculture, Athens, Georgia, USA (D.-H. Lee, D.E. Swayne)


[1] These authors contributed equally to this article.
Table. Time to most recent common ancestor for each gene segment of
genotype B.N6.1 influenza A(H5N6) viruses isolated in South Korea,
December 2017-January 2018 *

                 South Korea isolates, node 1

Gene            Mean                 95% HPD

PB2           2017 Sep             2017 Jul-Oct
PB1           2017 Sep             2017 Jul-Oct
PA            2017 Sep             2017 Aug-Oct
HA            2017 Sep             2017 Jul-Oct
NP            2017 Jul             2017 Apr-Sep
NA            2017 Jul             2017 Mar-Sep
M             2017 Aug             2017 May-Oct
NS            2017 Jul             2017 Apr-Oct

                   South Korea and Europe
                     isolates, node 2

Gene            Mean                 95% HPD

PB2           2017 May          2016 Dec-2017 Sep
PB1           2017 May             2017 Feb-Aug
PA            2017 Jul             2017 Apr-Sep
HA            2017 May             2017 Feb-Jul
NP            2017 Mar          2016 Nov-2017 Jun
NA            2017 Feb          2016 Jul-2017 Jul
M             2017 May             2017 Jan-Aug
NS            2017 Mar          2016 Nov-2017 Jun

                South Korea, Europe, Japan, Taiwan,
                  and Greece isolates, node 3

Gene            Mean                 95% HPD

PB2           2016 Mar          2015 Oct-2016 Jun
PB1           2016 Jul             2016 May-Aug
PA            2016 Oct             2016 Jul-Dec
HA            2016 Mar          2015 Dec-2016 May
NP            2016 Jan          2015 Jul-2016 May
NA            2015 Sep          2014 Aug-2016 Aug
M             2016 Mar             2016 Jan-May
NS            2016 Feb          2015 Oct-2016 Jun

* Nodes of the temporally structured maximum clade credibility
phylogenetic tree (online Technical Appendix Figure 4,
HA, hemagglutinin; HPD, highest posterior density; M, matrix;
NA, neuraminidase; NP, nucleoprotein; NS,
nonstructural; PA, polymerase acidic; PB, polymerase basic.
COPYRIGHT 2018 U.S. National Center for Infectious Diseases
No portion of this article can be reproduced without the express written permission from the copyright holder.
Copyright 2018 Gale, Cengage Learning. All rights reserved.

Article Details
Printer friendly Cite/link Email Feedback
Author:Kwon, Jung-Hoon; Jeong, Sol; Lee, Dong-Hun; Swayne, David E.; Kim, Yu-jin; Lee, Sun-hak; Noh, Jin-Yo
Publication:Emerging Infectious Diseases
Article Type:Report
Geographic Code:9SOUT
Date:Oct 1, 2018
Previous Article:Introduction of Eurasian-Origin Influenza A(H8N4) Virus into North America by Migratory Birds.
Next Article:Clinical Isolation and Identification of Haematospirillum jordaniae.

Terms of use | Privacy policy | Copyright © 2021 Farlex, Inc. | Feedback | For webmasters