Reassortant Clade 2.3.4.4 of Highly Pathogenic Avian Influenza A(H5N6) Virus, Taiwan, 2017

A highly pathogenic avian influenza A(H5N6) virus of clade 2.3.4.4 was detected in a domestic duck found dead in Taiwan during February 2017. The endemic situation and continued evolution of various reassortant highly pathogenic avian influenza viruses in Taiwan warrant concern about further reassortment and a fifth wave of intercontinental spread.

A highly pathogenic avian influenza A(H5N6) virus of clade 2.3.4.4 was detected in a domestic duck found dead in Taiwan during February 2017. The endemic situation and continued evolution of various reassortant highly pathogenic avian influenza viruses in Taiwan warrant concern about further reassortment and a fifth wave of intercontinental spread.
We report HPAIV H5N6 detection from a meattype duck in Taiwan in February 2017. One dead young Pekin-type domestic duck was found on a country road near the Xiuguluan River in Hualien County during wild bird and habitat surveillance for HPAIV by the Wild Bird Society of Taipei; the carcass was forwarded to the national laboratory of the Animal Health Research Institute (online Technical Appendix 1 Figure 1, https://wwwnc. cdc.gov/EID/article/24/6/17-2071-Techapp1.pdf).
We conducted complete genome sequencing and comparative phylogenetic analysis of the detected virus, A/duck/ Taiwan/1702004/2017(H5N6) (Dk/Tw/17), to trace the origin and understand its genetic features.
We detected Dk/Tw/17 virus by using reverse transcription PCR and isolated the virus by using egg inoculation as described previously (9). We conducted an intravenous pathogenicity index test according to the World Organisation for Animal Health Manual of Diagnostic Tests and Vaccines for Terrestrial Animals (http://www.oie. int/en/international-standard-setting/terrestrial-manual). We performed full-length genome sequencing by using reverse transcription PCR amplification and Sanger sequencing (9). We estimated maximum-likelihood phylogenies by using RAxML (10) and constructed a median-joining phylogenetic network of the hemagglutinin gene by using NETWORK 5.0 (online Technical Appendix).
We classified Dk/Tw/17 as an HPAIV on the basis of the amino acid sequence at the hemagglutinin cleavage site (PLRERRRKR/G) and its high lethality in chickens (intravenous pathogenicity index 3.0). Necropsy and histologic examination revealed virus-specific necrotic and inflammatory lesions in the pancreas, heart, and brain (online Technical Appendix 1 Figure 2). Phylogenetic analyses suggested that the Dk/Tw/17 virus belongs to clade 2.  Table). These viruses consistently clustered together with high bootstrap value (>70) in maximum-likelihood phylogenies across all 8 gene segments (online Technical Appendix Figures 3-10).
The genotype C5 comprises 17 H5N6 HPAIVs identified from wild waterfowl in China, Japan, and South Korea during November-December 2016; a virus identified from a chicken farm (A/chicken/Korea/H23/2016 [H5N6]) in South Korea in November 2016; and the Dk/Tw/17 virus. Genotype C5 is phylogenetically distinct from viruses that caused outbreaks in poultry farms in Japan and South

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Korea during 2016-2017. This genotype has independently evolved and been maintained in wild bird populations in the bird flyway of East Asia, highlighting how wild waterfowl play an important role in the maintenance and dissemination of this HPAIV. In addition, the median-joining phylogenetic network analysis suggests that the A/chicken/ Korea/H23/2016 (H5N6) is not the direct ancestor of the Dk/Tw/17 virus, which was likely caused by separate introduction from wild birds (Figure, panel B).
The site where the dead duck was collected is adjacent to a river and located near many ponds used for duck farming. After identification of Dk/Tw/17, intensified active surveillance conducted over 3 months detected additional clade 2.3.4.4C H5N6 HPAIVs from 12 farms in 4 counties (online Technical Appendix 1 Figure 1). Clade 2.3.4.4A H5Nx HPAIVs, mainly H5N2 and H5N8, have caused outbreaks in the poultry industry of Taiwan since January 2015 (9). In 2017, clade 2.3.4.4A H5Nx and 2.3.4.4C H5N6 HPAIVs were detected in domestic poultry. The endemic situation and continued evolution of various reassortant HPAIVs in domestic poultry warrants concern about further reassortment. Enhanced active surveillance in domestic and wild waterfowl is required to monitor the spread and onward reassortment in Taiwan and to inform the design of improved prevention and control strategies. H uman monkeypox is a rare zoonotic infection caused by an orthopoxvirus and characterized by smallpoxlike signs and symptoms (1). The disease is endemic to the Democratic Republic of the Congo. Reported outbreaks have occurred mainly in rural rainforest areas of the Congo basin and West Africa, caused by the Central and West African clades of the virus, respectively (1-6). The West African clade is associated with milder disease, fewer deaths, and limited human-to-human transmission. Since 1970, only ≈10 cases in West Africa had been reported; in 2003, a total of 81 cases (41% laboratory confirmed) were reported in the United States (2,7,8). In Nigeria, a case of human monkeypox in a 4-year-old child in the southeastern part