Investigation of Avian Influenza Infections in Wild Birds,Poultry and Humans in Eastern Dongting Lake,China

We investigated avian influenza infections in wild birds, poultry, and humans at Eastern Dongting Lake, China. We analyzed 6,621 environmental samples, including fresh fecal and water samples, from wild birds and domestic ducks that were collected from the Eastern Dongting Lake area from November 2011 to April 2012. We also conducted two cross-sectional serological studies in November 2011 and April 2012, with 1,050 serum samples collected from people exposed to wild birds and/or domestic ducks. Environmental samples were tested for the presence of avian influenza virus (AIV) using quantitative PCR assays and virus isolation techniques. Hemagglutination inhibition assays were used to detect antibodies against AIV H5N1, and microneutralization assays were used to confirm these results. Among the environmental samples from wild birds and domestic ducks, AIV prevalence was 5.19 and 5.32%, respectively. We isolated 39 and 5 AIVs from the fecal samples of wild birds and domestic ducks, respectively. Our analysis indicated 12 subtypes of AIV were present, suggesting that wild birds in the Eastern Dongting Lake area carried a diverse array of AIVs with low pathogenicity. We were unable to detect any antibodies against AIV H5N1 in humans, suggesting that human infection with H5N1 was rare in this region.     

Influenza a viruses from wild birds in Guatemala belong to the North American lineage

The role wild bird species play in the transmission and ecology of avian influenza virus (AIV) is well established; however, there are significant gaps in our understanding of the worldwide distribution of these viruses, specifically about the prevalence and/or significance of AIV in Central and South America. As part of an assessment of the ecology of AIV in Guatemala, we conducted active surveillance in wild birds on the Pacific and Atlantic coasts. Cloacal and tracheal swab samples taken from resident and migratory wild birds were collected from February 2007 to January 2010.1913 samples were collected and virus was detected by real time RT-PCR (rRT-PCR) in 28 swab samples from ducks (Anas discors). Virus isolation was attempted for these positive samples, and 15 isolates were obtained from the migratory duck species Blue-winged teal. The subtypes identified included H7N9, H11N2, H3N8, H5N3, H8N4, and H5N4. Phylogenetic analysis of the viral sequences revealed that AIV isolates are highly similar to viruses from the North American lineage suggesting that bird migration dictates the ecology of these viruses in the Guatemalan bird population.     

Molecular Characterization of Subtype H11N9 Avian Influenza Virus Isolated from Shorebirds in Brazil

Migratory aquatic birds play an important role in the maintenance and spread of avian influenza viruses (AIV). Many species of aquatic migratory birds tend to use similar migration routes, also known as flyways, which serve as important circuits for the dissemination of AIV. In recent years there has been extensive surveillance of the virus in aquatic birds in the Northern Hemisphere; however in contrast only a few studies have been attempted to detect AIV in wild birds in South America. There are major flyways connecting South America to Central and North America, whereas avian migration routes between South America and the remaining continents are uncommon. As a result, it has been hypothesized that South American AIV strains would be most closely related to the strains from North America than to those from other regions in the world. We characterized the full genome of three AIV subtype H11N9 isolates obtained from ruddy turnstones (Arenaria interpres) on the Amazon coast of Brazil. For all gene segments, all three strains consistently clustered together within evolutionary lineages of AIV that had been previously described from aquatic birds in North America. In particular, the H11N9 isolates were remarkably closely related to AIV strains from shorebirds sampled at the Delaware Bay region, on the Northeastern coast of the USA, more than 5000 km away from where the isolates were retrieved. Additionally, there was also evidence of genetic similarity to AIV strains from ducks and teals from interior USA and Canada. These findings corroborate that migratory flyways of aquatic birds play an important role in determining the genetic structure of AIV in the Western hemisphere, with a strong epidemiological connectivity between North and South America.     

Determining the Phylogenetic and Phylogeographic Origin of Highly Pathogenic Avian Influenza (H7N3) in Mexico

Highly pathogenic (HP) avian influenza virus (AIV) H7N3 outbreaks occurred 3 times in the Americas in the past 10 years and caused severe economic loss in the affected regions. In June/July 2012, new HP H7N3 outbreaks occurred at commercial farms in Jalisco, Mexico. Outbreaks continued to be identified in neighbouring states in Mexico till August 2013. To explore the origin of this outbreak, time resolved phylogenetic trees were generated from the eight segments of full-length AIV sequences in North America using BEAST. Location, subtype, avian host species and pathogenicity were modelled as discrete traits upon the trees using continuous time Markov chains. A further joint analysis among segments was performed using a hierarchical phylogenetic model (HPM) which allowed trait rates (location, subtype, host species) to be jointly inferred across different segments. The complete spatial diffusion process was visualised through virtual globe software. Our result indicated the Mexico HP H7N3 originated from the large North America low pathogenicity AIV pool through complicated reassortment events. Different segments were contributed by wild waterfowl from different N. American flyways. Five of the eight segments (HA, NA, NP, M, NS) were introduced from wild birds migrating along the central North American flyway, and PB2, PB1 and PA were introduced via the western North American flyway. These results highlight a potential role for Mexico as a hotspot of virus reassortment as it is where wild birds from different migration routes mix during the winter.     

Characterization of Avian-like Influenza A (H4N6) Virus Isolated from Caspian Seal in 2012

正Dear Editor Marine mammals are widely distributed and can be found almost in all coastal waters and coastlines around the world. The interface areas between marine and terrestrial environments provide natural habitats for aquatic and semiaquatic mammals as well as for reservoir species of avian influenza viruses (AIV)(Runstadler et al. 2013). Previous     

Are Poultry or Wild Birds the Main Reservoirs for Avian Influenza in Bangladesh?

Avian influenza viruses (AIV) are of great socioeconomic and health concern, notably in Southeast Asia where highly pathogenic strains, such as highly pathogenic avian influenza (HPAI) H5N1 and other H5 and H7 AIVs, continue to occur. Wild bird migrants are often implicated in the maintenance and spread of AIV. However, little systematic surveillance of wild birds has been conducted in Southeast Asia to evaluate whether the prevalence of AIV in wild birds is higher than in other parts of the world where HPAI outbreaks occur less frequently. Across Bangladesh, we randomly sampled a total of 3585 wild and domestic birds to assess the prevalence of AIV and antibodies against AIV and compared these with prevalence levels found in other endemic and non-endemic countries. Our study showed that both resident and migratory wild birds in Bangladesh do not have a particularly elevated AIV prevalence and AIV sero-prevalence compared to wild birds from regions in the world where H5N1 is not endemic and fewer AIV outbreaks in poultry occur. Like elsewhere, notably wild birds of the orders Anseriformes were identified as the main wild bird reservoir, although we found exceptionally high sero-prevalence in one representative of the order Passeriformes, the house crow (Corvus splendens), importantly living on offal from live bird markets. This finding, together with high sero- and viral prevalence levels of AIV in domestic birds, suggests that wild birds are not at the base of the perpetuation of AIV problems in the local poultry sector, but may easily become victim to AIV spill back from poultry into some species of wild birds, potentially assisting in further spread of the virus.     

Complete genome sequence of an H7N3 avian influenza virus isolated from ducks in southern China

We report here the complete genomic sequence of an H7N3 avian influenza virus (AIV) isolate, which was obtained from duck in 1996. This is the first report of this subtype of AIV being isolated from duck in Guangdong of Southern China. Genomic sequence and phylogenetic analyses showed that it was highly homologous with the wild bird virus A/ruddy turnstone/Delaware Bay/135/1996 (H7N3) and that all eight genes of this virus belonged to the North America gene pool. The availability of genome sequences is helpful to further investigations of epidemiology and evolution of AIV between waterfowl and wild birds.     

三江自然保护区野生迁徙水禽携带禽流感病毒和新城疫病毒状况的监测

[目的]为了对途经三江保护区的野生迁徙水禽携带禽流感病毒(AIV)和新城疫病毒(NDV)的状况进行有效监测.[方法]在2005年10月、2006年4月、2006年10月3个候鸟的迁徙季节从三江保护区采集了158只野鸟的咽拭子和肛拭子样本.应用SPF鸡胚盲传、血凝和血凝抑制试验和RT-PCR等方法进行了病毒的分离和鉴定.[结果]结果共分离到20株AIV和13株NDV.20株AIV均来自2006年10月采集的样品,经常规血清学分型鉴定分为12个亚型,11个亚型来源于绿头鸭,分别为H2N2(2/20),H2N6(2/20),H3N4(1/20),H3N6(2/20),H3N7(2/20),H3N8(2/20),H6N2(2/20),H11N2(1/20),H11N3(1/20),H11N5(2/20),H11N6(1/20),另外一株来源于白眉鸭,为H5N2(1/20).13株NDV则来自3个迁徙季节的5种不同水禽采,其中包括绿头鸭(8/13),豆雁(1/13),白额雁(1/13),绿翅鸭(1/13)和鸳鸯(2/13).[结论]这一结果表明,拥有极大种群数量、在世界范围内广泛分布的绿头鸭,被认为可能是AIV和NDV最重要的自然宿主之一,并在病毒的传播上比其他野生鸟类具有更为重要的生态学意义.     

渤海湾两株H2亚型禽流感病毒的 遗传进化分析

野鸟是禽流感病毒的自然储存库,病毒可以随着宿主的迁徙传播给其他野鸟与家禽。渤海湾是鸟类南北迁徙的重要停歇地,也是东亚-澳大利西亚鸟类迁徙通道的重要组成部分,每年有大量水鸟在渤海湾停歇,促进了禽流感病毒的传播。为了解渤海湾地区禽流感病毒的传播及进化与水鸟迁徙的相关性,2018年春季鸟类迁徙期间的4和5月份,在渤海湾采集鸻鹬类粪便样品2 120份,对样品进行检测,分离出2株H2亚型禽流感病毒。对这2株H2亚型禽流感病毒进行了分子特征及遗传进化分析,并结合渤海湾水鸟的环志回收数据,对H2亚型病毒的重组及遗传进化与水鸟迁徙的联系进行了分析。结果表明,2株分离株的HA蛋白裂解位点符合低致病性禽流感病毒的分子特征,它们的8个基因片段同源性均不高,其中879-H2N7的8个基因片段分别与我国福建和澳大利亚的毒株同源性最高,遗传关系最近;854-H2N8的8个基因片段分别与我国湖南以及日本、韩国、孟加拉国和越南的毒株同源性最高,遗传关系最近。渤海湾水鸟的环志回收数据分析表明,879-H2N7随着野鸟的迁徙在渤海湾、福建沿海和澳大利亚之间进行传播与扩散;854-H2N8可能跨越东亚-澳大利西亚和中亚-印度两条通道之间进行基因重组和进化,并会随着鸟类迁徙进行传播和扩散。     

Genetic Characterization of a Novel Recombinant H5N2 Avian Influenza Virus Isolated from Chickens in Tibet

In this report, a novel H5N2 avian influenza virus (AIV) was isolated from chickens in Tibet in 2010, western China. Phylogenetic analysis demonstrated that it was a natural reassortant between H9N2 and H5N1 subtypes. It is of note that this virus has an HP genotype with HA, PB2, M, and NS genes homologous to those of A/peregrine falcon/Hong Kong/2142/2008(H5N1)-like HPAIV isolated from dead wild birds. Publishing this genome information will contribute to the investigation of avian influenza epidemiology and to further research of AIV''s biological properties.     

西洞庭湖湿地越冬期野生植物多样性

本研究采用定点取样法,于2008～2009年越冬期对西洞庭湖湿地野生植物多样性进行了调查统计分析.结果表明:西洞庭湖湿地野生植物种类共73种,隶属29科,植物种类丰富,其中以菊科、十字花科野生植物为主.西洞庭湖区野生植物群落具有较大的物种丰富度,分布均匀,且不同种之间相遇几率较大,具有较高的物种多样性.为保护西洞庭湖区野生植物多样性提供了科学依据.     

H5N1 virus-like particle vaccine elicits cross-reactive neutralizing antibodies that preferentially bind to the oligomeric form of influenza virus hemagglutinin in humans

Transmission of pathogenic avian influenza viruses (AIV) from wild birds to domestic poultry and humans is continuing in multiple countries around the world. In preparation for a potential AIV pandemic, multiple vaccine candidates are under development. In the case of H5N1 AIV, a clear shift in transmission from clade 1 to clade 2 viruses occurred in recent years. The virus-like particle (VLP) represents an economical approach to pandemic vaccine development. In the current study, we evaluated the humoral immune response in humans vaccinated with H5N1 A/Indonesia/05/2005 (clade 2.1) VLP vaccine manufactured in Sf9 insect cells. The VLPs were comprised of the influenza virus hemagglutinin (HA), neuraminidase (NA), and matrix 1 (M1) proteins. In an FDA-approved phase I/II human clinical study, two doses of H5N1 VLPs at 15, 45, or 90 μg HA/dose resulted in seroconversion and production of functional antibodies. Moreover, cross-reactivity against other clade 2 subtypes was demonstrated using virus neutralization assays. H5N1 whole-genome fragment phage display libraries (GFPDL) were used to elucidate the antibody epitope repertoire in postvaccination human sera. Diverse epitopes in HA1/HA2 and NA were recognized by postvaccination sera from the two high-dose groups, including large segments spanning the HA1 receptor binding domain. Importantly, the vaccine elicited sera that preferentially bound to an oligomeric form of recombinant HA1 compared with monomeric HA1. The oligomeric/monomeric HA1 binding ratios of the sera correlated with the virus neutralizing titers. Additionally, the two high-dose VLP vaccine groups generated NA-inhibiting antibodies that were associated with binding to a C-terminal epitope close to the sialic acid binding site. These findings represent the first report describing the quality of the antibody responses in humans following AIV VLP immunization and support further development of such vaccines against emerging influenza virus strains.     

2004年初广东省高致病性禽流感(H5N1)疫点野生鸟类禽流感研究简报

2004年初禽流感疫情期间,针对广东省疫点地区开展了野生鸟类禽流感病毒（H5、H7、H9）调查取样工作,共获得血样76份、拭子125份、器官样21份。血清检测采用琼脂扩散（AGP）和血凝抑制试验（H1）,病毒检测用鸡胚法鉴定。分析结果：（1）只在非疫点的汕头南澳的八哥上检测到H9病毒;（2）雷州、罗定、海丰及汕头4个地区的血清检测结果中H5和H9阳性率分别为31．6％和23．7％;沿海地区阳性率极显著高于非沿海地区;（3）候鸟的阳性率极显著高于留鸟。因此,该次疫情中候鸟中存在传播禽流感病毒的可能性。     

H9N2禽流感病毒反向遗传系统转录／表达载体的构建和验证

设计带有BsmBI、BsaI或AarI酶切位点的引物,用RT-PCR扩增H9N2亚型禽流感病毒(AIV)的8个基因全长片段,克隆入双向转录/表达载体pHW2000,并在PB2、PB1和NA基因中共引入了3个沉默突变标签。将其2个表面基因(HA和NA基因)加上任意1个内部基因,而其它5个内部基因来自A/WSN/33,进行了6种3 5组合形式的基因重排,把相应组合的转录/表达质粒共转染COS-1细胞,均产生了预期组合、有感染性的H9N2亚型流感病毒,表明亲缘关系遥远的流感病毒可以互相获取基因片段产生重组病毒,提示表面结构基因和单个内部基因不足以限制H9N2AIV在哺乳动物细胞上的宿主范围,同时也验证了构建的8个转录/表达载体均能有效工作,为进一步研究H9N2亚型AIV基因结构与功能、AIV与宿主之间的关系打下了基础。     

Development of a SYBR green real-time PCR method for rapid detection of sheep pox virus

Background

Virus subtype H13N2, A/mallard/Kr/SH38-45/2010 (H13N2), was first isolated from a mallard fecal sample in South Korea.

Results

Phylogenetic analysis of all eight viral genes revealed that this virus emerged by genetic mixing between Eurasian and North American gene pools, and possibly between wild ducks and gulls. The H13 and N2 surface genes clustered together in a group with Eurasian isolates from gulls and wild birds, respectively. The PB2, PA, NP, M and NS segments belonged to the Eurasian lineage, whereas the PB1 gene clustered in the North American lineage. Furthermore, they showed a bird-dependent pattern in phylogenetic analysis: the M gene was similar to subtype H13 viruses within gulls, whereas other segments were similar to avian influenza viruses of other subtypes from wild ducks.

Conclusions

The data suggests that the novel reassortant H13N2 virus isolated in South Korea might have emerged by genetic reassortment between intercontinental and interspecies transmission in wild birds.     

2004年初广东省高致病性禽流感(H5N疫点野生鸟类禽流感研究简报

2004年初禽流感疫情期间,针对广东省疫点地区开展了野生鸟类禽流感病毒(H5、H7、H9)调查取样工作,共获得血样76份、拭子125份、器官样21份。血清检测采用琼脂扩散(AGP)和血凝抑制试验(HI),病毒检测用鸡胚法鉴定。分析结果:(1)只在非疫点的汕头南澳的八哥上检测到H9病毒;(2)雷州、罗定、海丰及汕头4个地区的血清检测结果中H5和H9阳性率分别为31·6%和23·7%;沿海地区阳性率极显著高于非沿海地区;(3)候鸟的阳性率极显著高于留鸟。因此,该次疫情中候鸟中存在传播禽流感病毒的可能性。     

我国部分禽流感病毒H5N1之HA序列变异演化分析

从GenBank上获得我国人(Homo sapiens)、家禽和野鸟42株H5N1亚型禽流感病毒的HA基因核酸序列,利用DNAStar分析HA蛋白关键位点氨基酸残基的变化,比较HA基因核苷酸序列同源性,构建遗传进化树.探讨我国部分人、家禽和野鸟H5N1病毒基因的遗传进化关系.序列分析结果表明:禽流感病毒H5N1亚型的HA基因持续地发生着变异,但并非以均一速度进行,时间间隔愈长,核苷酸同源性愈低;我国同一地区或临近地区,当年或前后两年发生的人及家禽感染的禽流感病毒高度同源.推测我国部分人发生的禽流感可能是通过家禽感染的;候鸟的迁徙在传播病毒过程中所起的作用有待深入探讨.     

A robust tool highlights the influence of bird migration on influenza A virus evolution

One of the fundamental unknowns in the field of influenza biology is a panoramic understanding of the role wild birds play in the global maintenance and spread of influenza A viruses. Wild aquatic birds are considered a reservoir host for all lowly pathogenic avian influenza A viruses (AIV) and thus serve as a potential source of zoonotic AIV, such as Australasian‐origin H5N1 responsible for morbidity and mortality in both poultry and humans, as well as genes that may contribute to the emergence of pandemic viruses. Years of broad, in‐depth wild bird AIV surveillance have helped to decipher key observations and ideas regarding AIV evolution and viral ecology including the trending of viral lineages, patterns of gene flow within and between migratory flyways and the role of geographic boundaries in shaping viral evolution (Bahl et al. 2009 ; Lam et al. 2012 ). While these generally ‘virus‐centric’ studies have ultimately advanced our broader understanding of AIV dynamics, recent studies have been more host‐focused, directed at determining the potential impact of host behaviour on AIV, specifically, the influence of bird migration upon AIV maintenance and transmission. A large number of surveillance studies have taken place in Alaska, United States—a region where several global flyways overlap—with the aim of detecting the introduction of novel, Australasian‐origin highly pathogenic H5N1 AIV into North America. By targeting bird species with known migration habits, long‐distance migrators were determined to be involved in the intercontinental movement of individual AIV gene segments, but not entire viruses, between the Australasian and North American flyways (Koehler et al. 2008 ; Pearce et al. 2010 ). Yet, bird movement is not solely limited to long‐distance migration, and the relationship of resident or nonmigratory and intermediate‐distance migrant populations with AIV ecology has only recently been explored by Hill et al. ( 2012 ) in this issue of Molecular Ecology. Applying a uniquely refined, multidimensional approach, Hill et al. validate the innovative use of stable isotope assays for qualifying migration status of wild mallards within the Pacific flyway. The authors reveal that AIV prevalence and diversity did not differ in wintering mallard ducks with different migration strategies, and while migrant mallards do indeed introduce AIV, these viruses do not circulate as the predominant viruses in resident birds. On the other hand, resident mallards from more temperate regions act as reservoirs, possibly contributing to the unseasonal circulation and extended transmission period of AIV. This study highlights the impact of animal behaviour on shaping viral evolution, and the unique observations made will help inform prospective AIV surveillance efforts in wild birds.     

Prevalence of avian influenza viruses, <Emphasis Type="Italic">Mycobacterium avium</Emphasis>, and <Emphasis Type="Italic">Mycobacterium avium</Emphasis>, subsp. <Emphasis Type="Italic">paratuberculosis</Emphasis> in marsh-dwelling passerines in Slovakia, 2008

Prevalence of the infectious respiratory agens, avian influenza virus (AIV), Mycobacterium avium (M. avium), and Mycobacterium avium subspecies paratuberculosis (MAP), was studied in migratory marsh-dwelling passerines captured in the Parížske močiare wetlands in Western Slovakia during 2008. Surveillance of 650 birds revealed a lower prevalence of AIV in spring (13.6%) than in summer (17.5%). A total of 14 different subtypes were detected in samples obtained from birds captured during the spring, with the most prevalent subtypes being H8N3, H6N4, H11N6 and H12N6. Subtypes H12N6, H6N6 and H2N5 were predominant in passerines captured during summer months. In eight cases, different AIV infections were detected in the oropharyngeal and cloacal samples originating from a single bird (H1N1 and H8N3; H1N3 and H9N3; H2N3 and H12N6; H2N1 and H8N1; H4N2 and H9N6; H5N5 and H11N6; H6N4 and H11N6; H7N1 and H10N3 in the oropharynx and cloaca, respectively). M. avium was detected in 9.2% and 0.8% of marsh-dwelling passerines captured during spring and summer, respectively. Only two birds were co-infected with AIV and M. avium. All birds were negative for MAP.