Ji-Ye Kim

Novel Reassortant Influenza A(H5N8) Viruses, South Korea, 2014

To the Editor: Highly pathogenic avian influenza (HPAI) viruses have caused considerable economic losses to the poultry industry and poses potential threats to animal and human health (www.oie.int/en/ and www.who.int/en/). Since 2003, influenza A(H5N1) viruses with a hemagglutinin (HA) gene derived from A/goose/Guandong/1/96–like viruses have become endemic to 6 countries (Bangladesh, China, Egypt, India, Indonesia, and Vietnam) (1) (www.cdc.gov/). Furthermore, HPAI viruses with an H5 subtype continue to undergo substantial evolution because of extensive genetic divergence and reassortment between other subtypes of influenza viruses. Especially in China, novel subtypes of H5 HPAI virus, such as influenza A(H5N2), influenza A(H5N5), and influenza A(H5N8) viruses, were reported during 2009–2011 (2,3).

Highly pathogenic avian influenza virus (H5N8) in domestic poultry and its relationship with migratory birds in South Korea during 2014.

Highly pathogenic H5N8 avian influenza viruses (HPAIVs) were introduced into South Korea during 2014, thereby caused outbreaks in wild birds and poultry farms. During the 2014 outbreak, H5N8 HPAIVs were isolated from 38 wild birds and 200 poultry farms (up to May 8, 2014). To better understand the introduction of these viruses and their relationships with wild birds and poultry farm, we analyzed the genetic sequences and available epidemiological data related to the viruses. Genetic analysis of 37 viruses isolated from wild birds and poultry farms showed that all of the isolates belonged to clade 2.3.4.6 of the hemagglutinin (HA) gene, but comprised two distinct groups. During the initial stage of the outbreak, identical isolates from each group were found in wild birds and poultry farms near Donglim Reservoir, which is a resting site for migratory birds, thereby indicating that two types of H5N8 HPAIVs were introduced into the lake at the same time. Interestingly, the one group of H5N8 HPAIV predominated around Donglim Reservoir, and the predominant virus was dispersed by wild birds among the migratory bird habitats in the western region of South Korea as time passed, and it was also detected in nearby poultry farms. Furthermore, compared with the results of the annual AIV surveillance of captured wild birds, which has been performed since 2008, more HPAIVs were isolated and H5 sero-prevalence was also detected during the 2014 outbreak. Overall, our results strongly suggest that migratory birds played a key role in the introduction and spread of viruses during the initial stage of the 2014 outbreak.

A Novel Avian Paramyxovirus (Putative Serotype 15) Isolated from Wild Birds

In January 2014, a viral hemagglutinating agent named UPO216 was isolated from fecal droppings of wild birds at the UPO wetland in South Korea during an avian influenza surveillance program. Electron microscopy identified the UPO216 virus as an avian paramyxovirus (APMV). Pathogenicity tests and molecular pathotyping revealed that the virus was avirulent in chickens. The UPO216 virus was assigned to a serological group antigenically distinct from known serotypes of APMV (−1, −2, −3, −4, −6, −7, −8, and −9) by hemagglutination inhibition test, despite showing weak cross-reactivity with APMV-1 and APMV-9. The UPO216 virus RNA genome is 15,180 nucleotides (nts) in length, encodes 3′-N-P(V/W)-M-F-HN-L-5′ in that order, and shows unique genetic characteristics in terms of genomic composition and evolutionary divergence (0.43 or greater from known serotypes of APMV). Phylogenetic analysis revealed that the UPO216 occupies a branch separate from APMV-1, -9, -12, and -13. Serologic surveillance of wild birds (n = 880; 15 species, five Orders) detected UPO216-reactive antibodies in 4% (20/494) of serum samples taken from five species of wild duck belonging to the Order Anseriformes. In particular, UPO216-specific antibodies showing no cross-reaction with other serotypes of APMV were detected in four species: Eurasian teal (1/36), European wigeon (1/73), mallard (4/139), and Spot-Billed duck (1/137). These results indicate that the UPO216 virus has antigenically and genetically unique characteristics distinct from known serotypes of APMV and likely has been circulating widely in wild duck species of the Order Anseriformes. Thus, we propose the UPO216 isolate as a prototype strain of a novel APMV serotype (putative APMV-15).

Genetic evolution of H5 highly pathogenic avian influenza virus in domestic poultry in Vietnam between 2011 and 2013

In spite of highly pathogenic avian influenza H5N1 vaccination campaigns for domestic poultry, H5N1 viruses continue to circulate in Vietnam. To estimate the prevalence of avian influenza virus in Vietnam, surveillance was conducted between November 2011 and February 2013. Genetic analysis of 312 highly pathogenic avian influenza H5 viruses isolated from poultry in Vietnam was conducted and possible genetic relationships with strains from neighboring countries were investigated. As previously reported, phylogenetic analysis of the avian influenza virus revealed two H5N1 HPAI clades that were circulating in Vietnam. Clade 1.1, related to Cambodian strains, was predominant in the southern provinces, while clade 2.3.2.1 viruses were predominant in the northern and central provinces. Sequence analysis revealed evidence of active genetic evolution. In the gene constellation of clade 2.3.2.1, genotypes A, B, and B(II) existed during the 2011/2012 winter season. In June 2012, new genotype C emerged by reassortment between genotype A and genotype B(II), and this genotype was predominant in 2013 in the northern and central provinces. Interestingly, enzootic Vietnamese clade 2.3.2.1C H5 virus subsequently reassorted with N2, which originated from wild birds, to generate H5N2 highly pathogenic avian influenza, which was isolated from duck in the northeast region. This investigation indicated that H5N1 outbreaks persist in Vietnam and cause genetic reassortment with circulating viruses. It is necessary to strengthen active influenza surveillance to eradicate highly pathogenic avian influenza viruses and sever the link between highly pathogenic avian influenza and other circulating influenza viruses.

A Survey of Exposure Level and Lifestyle Factors for Perfluorooctanoate and Perfluorooctane Sulfonate in Human Plasma from Selected Residents in Korea

Following few decades of commercial use, perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS) have been found in human blood and serum. We determined the amounts of PFOA and PFOS in human plasma (n = 183) and the effects of multiple uses of food-contact materials and smoking habits and alcohol consumption using liquid chromatography time-of-flight mass spectrometry (LC/TOF-MS). For the paper cups, the PFOA level in the plasma of the heavy user group was 1.37 times higher than that of the light user group. However, no association between the effects of multiple uses of food-contact materials and the plasma levels of PFOA and PFOS was found, except for paper cups. Active smokers had lower plasma levels of PFOA and PFOS than non-smokers. We show that multiple uses of food-contact materials do not appear to be a significant source of PFOA and PFOS.

Genetic characterization and evolutionary analysis of Newcastle disease virus isolated from domestic duck in South Korea

Domestic ducks are considered a potential reservoir of Newcastle disease virus. In the study, a Newcastle disease virus (NDV) isolated from a domestic duck during surveillance in South Korea was characterized. The complete genome of the NDV isolate was sequenced, and the phylogenetic relationship to reference strains was studied. Phylogenetic analysis revealed that the strain clustered in genotype I of Class II ND viruses, has highly phylogenetic similarity to NDV strains isolated from waterfowl in China, but was distant from the viruses isolated in chickens and vaccine strains used in South Korea. Pathogenicity experiment in chickens revealed it to be a lentogenic virus. The deduced amino acid sequence of the cleavage site of the fusion (F) protein confirmed that the isolate contained the avirulent motif (112)GKQGRL(117) at the cleavage site and caused no apparent disease in chickens and ducks. With phylogeographic analysis based on fusion gene, we estimate the origin of an ancestral virus of the isolate and its sister strain located in China around 1998. It highlights the need of continuous surveillance to enhance current understanding of the molecular epidemiology and evolution of the pathogenic strains.

Efficient self-assembly and protective efficacy of infectious bursal disease virus-like particles by a recombinant baculovirus co-expressing precursor polyprotein and VP4.

BackgroundVirus-like particle (VLP) technology is considered one of the most promising approaches in animal vaccines, due to the intrinsic immunogenic properties as well as high safety profile of VLPs. In this study, we developed a VLP vaccine against infectious bursal disease virus (IBDV), which causes morbidity and mortality in chickens, by expressing a baculovirus in insect cells.MethodsTo improve the self-proteolytic processing of precursor polyprotein (PP), we constructed a recombinant baculovirus transfer vector that co-expresses PP and the VP4 protease gene of IBDV.ResultsExpression and VLP assembly of recombinant proteins and antigenicity of the VLP were examined by Western blotting, ELISA, and transmission electron microscopy. In animal experiments, vaccination with the recombinant VLP induced strong and uniform humoral immunity and provided complete protection against challenge with very virulent (vv) IBDV in SPF chickens (n = 12). As determined by the bursa of Fabricius (BF)/body weight (B/BW) ratio, the protection against post-challenge bursal atrophy was significantly higher (P < 0.001) in VLP-vaccinated birds than in non-vaccinated controls.ConclusionsSince the protective efficacy of the VLP vaccine was comparable to that of a commercially available inactivated vaccine, the recombinant VLP merits further investigation as an alternative means of protection against vvIBD.

Genetic Diversity of Avian Paramyxovirus Type 6 Isolated from Wild Ducks in the Republic of Korea

Abstract:  Eleven avian paramyxovirus type 6 (APMV-6) isolates from Eurasian Wigeon (n=5; Anas penelope), Mallards (n=2; Anas platyrhynchos), and unknown species of wild ducks (n=4) from Korea were analyzed based on the nucleotide (nt) and deduced amino acid sequences of the fusion (F) gene. Fecal samples were collected in 2010–14. Genotypes were assigned based on phylogenetic analyses. Our results revealed that APMV-6 could be classified into at least two distinct genotypes, G1 and G2. The open reading frame (ORF) of the G1 genotype was 1,668 nt in length, and the putative F0 cleavage site sequence was 113PAPEPRL119. The G2 genotype viruses included five isolates from Eurasian wigeons and four isolates from unknown waterfowl species, together with two reference APMV-6 strains from the Red-necked Stint (Calidris ruficollis) from Japan and an unknown duck from Italy. There was an N-truncated ORF (1,638 nt), due to an N-terminal truncation of 30 nt in the signal peptide region of the F gene, and the putative F0 cleavage site sequence was 103SIREPRL109. The genetic diversity and ecology of APMV-6 are discussed.

Seroprevalance of Newcastle Disease Virus in Wild Birds in Korea

Newcastle disease virus (NDV) infects a variety of birds with a wide range of clinical signs from asymptomatic to severe. During a 10-month period in 2011, a total of 1,024 sera from wild birds including 42 species of birds in 8 orders were collected and the seroprevalence of NDV in wild birds was evaluated by hemagglutination inhibition (HI) test. Evidence of NDV infection was observed in 12.6% (129/1,024) of wild birds with a maximum prevalence reported in Mandarin duck (27.8%, 32/115) followed by Mallard duck (20.8%, 57/274), Spot-billed duck (11.9%, 36/303), Pintail (2.9%, 1/34), Black-tailed gull (2.9%, 1/34), White-fronted goose (1.8%, 1/56) and Common teal (1.4%, 1/69). None of the other 35 species of wild birds were antibody-positive for NDV. Mandarin duck, Mallard duck and Spot-billed duck showed high sero-prevalance of 12.2% to 42% during winter season (November to March). Our results indicate that Mandarin duck, Mallard duck and Spot-billed duck might be natural reservoirs for NDV in Korea and the prevalence of NDV infection in wild birds displayed a seasonal pattern with high prevalence of NDV in winter season (November to March).

Biological Properties of Vero Cell-Adapted Newcastle Disease Virus

Newcastle disease virus (NDV) Kr005/V strain was generated through 55 serial passages of NDV Kr005 strain in Vero cells. The Kr005/V virus yielded high infective titers of 10 7.8 TCID50/mL in Vero cells and the infected cells showed cytopathic effects such as marked cell rounding, though less frequent syncytia. The Kr005/V virus was heat-stable and classified into the lentogenic type with a Mean Death Time (MDT) of 120h or greater while the Kr005 strain was heat-labile and velogenic (MDT of 49.6 h). Only the single amino acid substitution (T to S) was observed at position 433 of the HN protein of the Kr005/V strain, whereas no amino acid change was found in the F protein. The Kr005/V input virus correlated well (correlation coefficient r 2 =0.97) with the Kr005 virus when ten field sera were tested by virus neutralization test. The biolo-