Intestinal influenza: replication and characterization of influenza viruses in ducks.

Influenza A viruses isolated from the cloaca of naturally infected feral ducks replicate in the lungs and in the cells lining the intestinal tract of feral and domestic ducks. Despite the low pH of the gizzard, the duck influenza viruses reach the intestines via the digestive tract and are found in high concentration in the feces. The viruses retain infectivity in fecal material for at least 30 days at 4° and for 7 days at 20°. The morphology of one strain of intestinal duck influenza virus (Hav7 Neg2) that had never been passed in embryonated eggs and was isolated from the feces was roughly spherical and fairly uniform in size and shape. However, another strain of duck influenza virus studied (Hav3 Nav6) was predominantly filamentous, suggesting that the morphology of influenza viruses in their natural hosts varies from strain to strain. After passage in the chick embryo each strain retained the morphological characteristics found in the feces. In contrast to duck influenza viruses, representative human influenza viruses of the HON1, H3N2, and Hswl Nl subtypes replicate only in the upper respiratory tract of ducks. The duck influenza viruses are more stable to low pH than human strains and retain infectivity for over 30 days in nonchlorinated river water at 0° and for 4 days at 22°. The susceptibility of ducks to infection with human and avian strains of influenza virus and the possibility of transmission to animal species through the water supply suggests that ducks may be important in the ecology of influenza viruses. The possibility of “intestinal influenza” virus vaccines is considered.     

Isolation and characterization of influenza A viruses from wild ducks in northern Japan: appearance of HSW1 antigens in the Japanese duck population.

Twenty-six influenza A viruses were isolated from cloacal and tracheal samples of 235 resident and 396 migratory ducks in Miyagi prefecture, Japan, in 1977--78. Of these, twelve were antigenically related to the avian-origin HSW1 virus, A/duck/Alberta/35/76 (HSW1N1), but their neuraminidase antigens were characterized as Nav2-3, Nav4 or N2. These antigenic configuration have not previously been reported. In addition, one strain in which the neuraminidase antigen was identified as Nav4, was demonstrated to be a mixture of two haemagglutinins, HSW1 and Hav7. Two distinct strains were separated from the mixture and characterized as HSW1Nav4 and Hav7Nav4. The antigenic identification of an additional 13 influenza A viruses revealed the presence of six haemagglutinin subtypes (Hav1, Hav3, Hav4, Hav6, Hav7, and Hav8) and five neuaraminidase subtypes (Nav1, Nav2-3, Nav4, Neq2, and N2) in various combinations. The results suggest that the avian influenza A viruses among feral ducks may be isolated in various combinations of haemagglutinins and neuraminidase subtypes in Japan, and that feral ducks may be the site of genetic recombination occurring as a result of dual infection with different subtypes of influenza A virus.     

Properties of Hav6Neq2 and Hswl(H0)Hav2 influenza viruses isolated from waterfowl in southern Turkmenia

Three influenza A virus strains were isolated from shorebirds in October, 1977, in southern Turkmenia, in the vicinities of Tedzhen water reservoir. From a common tern, A/Sterna hirundo/Turkmenia/45/77 strain was isolated with the antigenic formula Hav6Neq2, from a teal and a black-headed gull influenza A/Anas crecca/Turkmenia/4/77 and A/Larus ridibundus/Turkmenia/13/77 strains with previously unknown combination of surface antigens Hswl(H0)Nav2 were recovered. By the molecular weight of the heavy (HA1 59,000 d) and light (HA2 24,000 d) chains of hemagglutinin, the Turkmenian viruses A/Larus ridibundus/Turkmenia/13/77 and A/Anas crecca/Turkmenia/4/77 are similar to each other and to the strains having H0 hemagglutinin: A/PR8/34 (H0N1) and A/Whale/PO/19/76 (H09Nav2). The Turkmenian viruses are characterized by a low content of the light hemagglutinin chain (HA2) which is typical of the viruses with Hsw1 hemagglutinin: A/New Jersey/8/76 (Hsw1N1) and A/SW/Wisk/68 (Hsw1N1).     

Genetic analysis of H3 subtype influenza viruses isolated from domestic ducks in northern China during 2004–2005

The broad distribution and prevalence of H3 subtype influenza viruses in avian and mammalian hosts constitutes a global threat to both human and veterinary health. In this present study, six H3N8 influenza viruses isolated from domestic ducks during 2004–2005 in northern China were genetically and phylogenetically characterized. Sequence analysis showed that HA, NA, and M genes of all the six H3N8 isolates had a close relationship with those of Equine/Jilin/1/89 (H3N8) virus, which once caused outbreak in equine populations in northern China. The PB2 and PA genes of the viruses possessed the highest similarities with highly pathogenic avian H5N1 influenza viruses currently circulating in this region. These findings emphasize the importance of avian influenza virus surveillance in this region for understanding the genesis and emergency of novel reassortants with pandemic potential.     

Molecular characterization and pathogenicity of swine influenza H9N2 subtype virus A/swine/HeBei/012/2008/(H9N2)

The H9N2 subtype influenza virus (IV) is a remarkable member of the influenza A viruses because it can infect not only chickens, ducks and pigs, but also humans. Pigs are susceptible to both human and avian influenza viruses and have been proposed to be intermediate hosts for the generation of pandemic influenza viruses through reassortment or adaptation to the mammalian host. To further understand the genetic characteristics and evolution, we investigated the source and molecular characteristics of the H9N2 subtype swine influenza virus (SIV), and observed its pathogenicity in BALB/c mice. The BALB/c mice were inoculated intranasally with 100 median mouse infectious dose of A/swine/HeBei/012/2008/(H9N2) viruses to observe the pathogenicity. The HA, NP, NA and M gene were cloned, sequenced and phylogenetically analyzed with related sequences available in GenBank. The infected mice presented with inactivity, weight loss and laboured respiration, while the pathological changes were characterized by diffuse alveolar damage in the lung. The nucleotide and deduced amino acid sequence of HA, NP, NA and M gene was similar with that of A/chicken/Hebei/4/2008(H9N2). The HA protein contained 6 glycosylation sites and the motif of HA cleavage site was PARSSR GLF, which is characteristic of low pathogenic IV. In the HA, NP, M and NA gene phylogenetic trees, the isolate clustered with A/chicken/Hebei/4/2008(H9N2). The isolate possibly came from A/chicken/Hebei/4/2008(H9N2) and was partially varied during its cross-species spread.     

Phylogenetic Analysis of Hemagglutinin and Neuraminidase Genes of H9N2 Viruses Isolated from Migratory Ducks

Genetic analysis indicated that the pandemic influenza strains derived from wild aquatic birds harbor viruses of 15 hemagglutinin (HA) and 9 neuraminidase (NA) antigenic subtypes. Surveillance studies have shown that H9N2 subtype viruses are worldwide in domestic poultry and could infect mammalian species, including humans. Here, we genetically analyzed the HA and NA genes of five H9N2 viruses isolated from the migratory ducks in Hokkaido, Japan, the flyway of migration from Siberia during 1997–2000. The results showed that HA and NA genes of these viruses belong to the same lineages, respectively. Compared with those of A/quail/Hong Kong/G1/97-like and A/duck/Hong Kong/Y280/97-like viruses, HA and NA of the migratory duck isolates had a close relationship with those of H9N2 viruses isolated from the chicken in Korea, indicating that the Korea H9N2 viruses might be derived from the migratory ducks. The NA genes of the five isolates were located in the same cluster as those of N2 viruses, which had caused a human pandemic in 1968, indicating that the NA genes of the previous pandemic strains are still circulating in waterfowl reservoirs. The present results further emphasize the importance of carrying out molecular epidemiological surveillance of H9N2 viruses in wild ducks to obtain more information for the future human influenza pandemics preparedness.     

New avian influenza A virus subtype combination H5N7 identified in Danish mallard ducks

During the past years increasing incidences of influenza A zoonosis have made it of uppermost importance to possess methods for rapid and precise identification and characterisation of influenza A viruses. We present here a convenient one-step RT-PCR method that will amplify full-length haemagglutinin (HA) and neuraminidase (NA) directly from clinical samples and from all known subtypes of influenza A. We applied the method on samples collected in September 2003 from a Danish flock of mallards with general health problems and by this a previously undescribed influenza A subtype combination, H5N7, was identified. The HA gene showed great sequence similarity to the highly pathogenic avian influenza A virus (HPAIV) A/Chicken/Italy/312/97 (H5N2); however, the cleavage site sequence between HA1 and HA2 had a motif typical for low pathogenic avian influenza viruses (LPAIV). The full-length NA sequence was most closely related to the HPAIV A/Chicken/Netherlands/01/03 (H7N7) that infected chickens and humans in the Netherlands in 2003. Ten persons with direct or indirect contact with the Danish mallard ducks showed signs of influenza-like illness 2-3 days following the killing of the ducks, but no evidence of influence infections was detected. To our knowledge this is the first report of an H5N7 influenza A virus.     

Characterization of a new avian-like influenza A virus from horses in China.

In March 1989 a severe outbreak of respiratory disease occurred in horses in the Jilin and Heilongjiang provinces of Northeast China that caused up to 20% mortality in some herds. An influenza virus of the H3N8 subtype was isolated from the infected animals and was antigenically and molecularly distinguishable from the equine 2 (H3N8) viruses currently circulating in the world. The reference strain A/Equine/Jilin/1/89 (H3N8) was most closely related to avian H3N8 influenza viruses. Sequence comparisons of the entire hemagglutinin (HA), nucleoprotein (NP), neuraminidase (NA), matrix (M), and NS genes along with partial sequences of the three polymerase (PB1, PB2, PA) genes suggest that six of the eight gene segments (PA, HA, NP, NA, M, NS) are closely related to avian influenza viruses. Since direct sequence analysis can only provide a crude measure of relationship, phylogenetic analysis was done on the sequence information. Phylogenetic analyses of the entire HA, NP, M, and NS genes and of partial sequences of PB1, PB2, and PA indicated that these genes are of recent avian origin. The NP gene segment is closely related to the gene segment found in the newly described H14 subtype isolated from ducks in the USSR. The A/Equine/Jilin/1/89 (H3N8) influenza virus failed to replicate in ducks, but did replicate and cause disease in mice on initial inoculation and on subsequent passaging caused 100% mortality. In ferrets, the virus caused severe influenza symptoms. A second outbreak of influenza in horses in Northeast China occurred in April 1990 in the Heilongjiang province with 48% morbidity and no mortality. The viruses isolated from this outbreak were antigenically indistinguishable from those in the 1989 outbreak and it is probable that the reduced mortality was due to the immune status of of the horses in the region. No influenza was detected in horses in Northern China in the spring, summer, or fall of 1991 and no influenza has been detected in horses in adjacent areas. Our analysis suggests that this new equine influenza virus in horses in Northeast China is the latest influenza virus in mammals to emerge from the avian gene pool in nature and that it may have spread to horses without reassortment. The appearance of this new equine virus in China emphasizes the potential for whole avian influenza viruses to successfully enter mammalian hosts and serves as a model and a warning for the appearance of new pandemic influenza viruses in humans.(ABSTRACT TRUNCATED AT 250 WORDS)     

Origin of the hemagglutinin gene of H3N2 influenza viruses from pigs in China

Influenza viruses of the H3N2 subtype similar to Aichi/2/68 and Victoria/3/75 persist in pigs many years after their antigenic counterparts have disappeared from humans (Shortridge et al. (1977). Science 19, 1454-1455). To provide information on the mechanism of conservation of these influenza viruses in pigs, the hemagglutinin (HA) of four isolates from swine derived from Taiwan and Southern China were analyzed antigenically and genetically. The reactivity pattern of these viruses with a panel of monoclonal antibodies indicates that the HAs of these swine viruses were antigenically closely related to duck H3 and early human H3 viruses. Sequence analysis of the H3 genes from three swine viruses revealed that the swine H3 genes are more closely related to the duck genes than to early human H3 virus (A/Aichi/2/68). The degree of sequence homology of these genes is extremely high (more than 96.5%). Furthermore, the deduced amino acid sequence of the three swine HAs at residues 226 to 228 in the proposed receptor-binding site is Gln-Ser-Gly and is common with the majority of avian influenza viruses. These findings indicate that these H3 viruses may have been introduced into pigs from ducks. The HA gene of the fourth swine influenza virus from Southern China was genetically equally related to avian and early human H3 strains although the sequence through the receptor-binding pocket (226-228) was typical of a human H3 virus, suggesting that either this swine HA gene was derived from ducks or an early human H3 virus was introduced into the pig population where the virus accumulated substantial mutations. The present strains revealed genetic heterogeneity of swine H3 influenza viruses in nature.     

Restricted infectivity of a human-Lineage H3N2 influenza A virus in pigs is hemagglutinin and neuraminidase gene dependent

Influenza A viruses cause pandemics at sporadic intervals. Pandemic viruses can potentially be introduced into the human population through in toto transfer of an avian influenza virus or through reassortment between avian and human strains. Pigs are believed to play a central role in the creation of pandemic viruses through reassortment because of their susceptibility to infection with both avian and human influenza viruses. However, we recently found that a human-lineage H3N2 influenza virus was highly restricted in its ability to infect pigs after intranasal inoculation. We hypothesized that this restricted infectivity phenotype was controlled by the hemagglutinin (HA) and neuraminidase (NA). To test this, we infected pigs with reverse genetics-created HA plus NA reassortant viruses. Specifically, introduction of the HA and NA genes of a contemporary H3N2 swine virus into the genetic background of the wholly human virus resulted in a significant increase in virus shedding and pathogenicity. These data indicate that the HA/NA can play important roles in controlling human influenza virus infectivity in pigs. The results further support the premise that a barrier exists to human influenza virus infection in pigs, which may limit the role of pigs in pandemic virus creation through reassortment of human and avian influenza viruses.     

Sequence and phylogenetic analysis of H2N7 avian influenza viruses isolated from domestic ducks in Zhejiang Province,Eastern China, 2013

Two H2N7 avian influenza viruses (AIVs) were isolated from domestic ducks in live poultry markets in Zhejiang Province, Eastern China, 2013. All viruses were characterized by whole-genome sequencing with subsequent phylogenetic analysis and genetic comparison. Phylogenetic analysis of all eight viral genes showed that the viruses clustered in the Eurasian lineage of AIVs and originated from genes reassortment among different viruses co-circulating in domestic ducks in Eastern China. The hemagglutinin cleavage site of all viruses indicated that the two strains were low-pathogenic avian influenza viruses. Considering the important role of the domestic ducks in the dissemination and reassortment of AIVs, continued surveillance of circulating H2 subtype AIVs in domestic ducks in live poultry markets is needed.     

猪型(H1N1)流感病毒血凝素和神经氨酸酶基因来源的研究

目的 研究2002年我国内地从猪群中分离的猪型(H1N1)毒株HA和NA基因来源。及其使猪致病的原因。方法 用PCR扩增目的基因，用P^GEM-T Easy Vector，4℃过夜连接，重组质粒转入DH-10B细菌，筛选阳性菌落，酶切鉴定，送六合通公司自动测序，并作进化树分析。结果 3株猪型(H1N1)病毒的HA和NA基因属猪型(H1N1)流感病毒，而不同于其他禽或人的H1N1亚型流感病毒。2002年猪型毒株由1991年猪型毒株演变而来。近来我国内地猪群中猪型毒株活动增强，其对猪能致病是由于病毒粒HA和NA蛋白抗原性发生变异所造成。结论 3株猪型病毒的HA和NA基因来源于猪型(H1N1)毒株。近来猪型毒株对猪具有致病性和活动增强是由于其HA和NA蛋白分子上氨基酸序列发生替换所造成。     

Human influenza a viral genes responsible for the restriction of its replication in duck intestine

Although influenza A viruses are occasionally transmitted from one animal species to another, their host range tends to be restricted. Currently circulating human influenza A viruses are thought to have originated from avian viruses, yet none of these strains replicate in duck intestine, a major site of avian virus replication. Although the hemagglutinin (HA) and neuraminidase (NA) genes are known to restrict human virus replication in ducks, the contribution of the other viral genes remains unknown. To determine the genetic basis for host range restriction of the replication of human influenza A virus in duck intestine, we first established a reverse genetics system for generating A/Memphis/8/88 (H3N2) (Mem/88) and A/mallard/New York/6750/78 (H2N2) (Mal/NY) viruses from cloned cDNAs. Using this system, we then attempted to generate reassortant viruses with various combinations of candidate genes. We were able to generate single-gene reassortants, which possessed PB2, NP, M, or NS from Mem/88, with the remainder from Mal/NY. Despite unsuccessful production of other single-gene reassortants from Mem/88, we did generate reassortant viruses comprised of both the HA and the NA, all three polymerase genes (PB2, PB1, and PA), or all polymerase genes and the NP gene from Mem/88, with the rest derived from Mal/NY. Among these reassortants, only those possessing the M or NS gene from Mem/88 and the remainder from Mal/NY replicated in duck intestine. These results indicate incompatibility between the genes of avian and human influenza A viruses and indicate that all genes other than the M and NS restrict replication of human influenza A virus in duck intestine.     

我国猪群中H9N2亚型毒株HA和NA基因特性的研究

目的 了解我国内地从猪中分离到H9N2亚型毒株HA和NA基因来源及它们使猪致病的原因。方法 用PCR扩增目的基因，与P^GEM-T Easy Vector4℃过夜连接，重组质粒转化DH-10β细菌，筛选阳性菌落，酶切鉴定，测序。然后，进行进化树分析。结果 两株猪H9N2毒株HA蛋白分子上第226位上氨基酸为L，这与从人和猪所分离出的H9N2毒株相同，其连接肽属对禽致病的毒株，但它们的序列为R-L-S-R，而不是R-S-S-R；其NA蛋白茎区第62～64位存在掉失，这与A／Shaoguarn/408／98，A／Swine／Hong Kong／9／98及A／Duck／Hong Kong／y280／97(H9N2)毒株相同；HA与NA基因进化树分析表明，两株猪H9N2毒株的HA基因接近于A／Chicken／Hong Kong／G23／97和A／Chicken／Hong Kong／G9／97．而NA基因接近于A／Shaoguan／408／98毒株。结论 两株猪H9N2亚型毒株的HA和NA基因可能性最大来自禽H9N2毒株。由于其HA蛋白分子上连接肽氨基酸序列发生替换，可能造成了它们对猪具有致病性。禽H9N2毒株NA蛋白茎区氨基酸掉失，造成了它们能直接感染猪。     

Characterization of an H4N2 avian influenza virus isolated from domestic duck in Dongting Lake wetland in 2009

In January 2009, an H4N2 subtype of avian influenza virus [A/duck/Hunan/8-19/2009 (H4N2)] was isolated from domestic ducks in Dongting Lake wetland. The whole genome of the virus was sequenced and the results indicated that multiple gene segments of the virus had a high homology with viruses isolated from wild waterfowl, which indicated that the virus was probably transmitted from wild waterfowl to domestic ducks. Phylogenetic analysis revealed that the each gene belonged to the Eurasian lineage of avian influenza viruses, but genetic reassortment occurs between viruses of different subtypes.     

Further evidence of the circulation of PMV-4 and influenza viruses with N2-1957 enzyme in the migratory waterfowls

In the years 1980-1984, one paramyxovirus type 4 and 11 influenza viruses were isolated from cloacal swabs collected from migratory waterfowls in Fed. Rep. Germany. One influenza virus of H4N8 subtype was isolated from swabs of commercial ducks collected at an abbatoir. Seven of 10 influenza strains, isolated from mallard ducks and coot were identified as a mixture of 2-3 strains of H1, H4, and H5 subtype; 3 virus strains from the same locality relate antigenically to subtype H4 with enzyme serologically identical with N2--Singapore/57 as demonstrated by means of polyclonal and monoclonal antibody.     

Influenza virus subtypes in aquatic birds of eastern Germany

Summary We report the findings of a 12-year surveillance study (1977–89) of avian influenza A viruses in eastern Germany. Viruses were isolated directly from feral ducks (n=236) and other wild birds (n=89); from domestic ducks (n=735) living on a single farm; and from white Pekin ducks (n=193) used as sentinels for populations of wild aquatic birds; mainly sea birds. The efficiency of virus isolation was 9.9% overall, with considerable variability noted among species: 8.7% in wild ducks, 0.9% in other feral birds and 38% in Pekin ducks. Use of sentinel ducks in wild pelagic bird colonies improved virus detection rates fivefold, suggesting that this approach is advantageous in ecological studies. Among the 40 different combinations of hemagglutinin (HA) and neuraminidase (NA) subtypes we identified, H6N1 predominated (23.6% for all avian species), followed by H4N6 (11%). Among individual species, the frequency profiles favored H2N3 (20.8%) and H4N6 (20.3%) in feral ducks; H7N7 (22.3%), H4N6 (24.4%) and H2N3 (10.4%) in Pekin ducks used as sentinels; and H6N1 (34.8%) and H6N6 (15.1%) in domestic ducks maintained on a single farm. By relying on sentinel birds for serological assays, it was possible to trace an influenza season in feral swan populations, beginning in August and continuing through the winter months. Comparison of subtype distribution of influenza viruses for Europe and North American showed significant differences. This supports the fact of two geographically distinct gene pools of influenza viruses in birds connected with their distinct flyways of each hemisphere. The high frequency of isolation of H2 influenza viruses is of considerable interest to those interested in the recycling of this subtype in humans. Similarly the frequent isolation of H7N7 influenza viruses raises concern about reservoirs of potentially pathogenic influenza virus for domestic poultry. Our results confirm the existence of a vast reservoir of influenza A viruses in European aquatic birds, which possesses sufficient diversity to account for strains that infect lower animals and humans.This article is dedicated to the memory of Dr. Herbert Sinnecker who died in 1991 at the age of 61. He was the Director of the Institute of Viral Zoonosis in the former German Democratic Republic (GDR). It was Herbert Sinnecker's forsight and understanding of the need to resolve the origin of human and animal influenza pandemics that initiated the studies described in this article. He developed novel epidemiological and ecological methods that permitted definition of the influenza virus gene pool in central Europe. The unification of Germany made it possible to publish this article; otherwise, the studies encouraged and organized by H. Sinnecker would have been lost to the scientific community.     

Novel influenza A viruses isolated from Canadian feral ducks: including strains antigenically related to swine influenza (Hsw1N1) viruses.

Twelve influenza A viruses, antigenically related to the Ho, H1 and Hsw1 subtypes, were isolated from cloacal samples of feral ducks in Canada. Antigenic comparisons showed that these viruses were most closely related to the recent HSW1N1 isolates from man and pigs, whereas in vivo pathogenicity tests revealed differences between the Hsw1N1 viruses from the ducks and those from humans and pigs. Antigenic characterization of 94 additional influenza A viruses from the ducks showed four haemagglutinin subtypes (Hav1, Hav4, Hav5 and Hav7), an unclassified haemagglutinin, and six neuraminidase subtypes (N1, N2, Neq2, Nav1, Nav2 and Nav5) in various combinations, some of which are novel and have not previously been reported. Three of these duck influenza viruses possessed a haemagglutinin antigenically related to that of classical fowl plaque virus. A much higher percentage of virus isolations were from juvenile ducks (18.5%) than from adults (5%). All of the ducks, from which viruses were isolated, appeared healthy at the time of sampling. Serological studies on a limited number of humans and domestic birds living in close proximity to the Canadian ducks revealed no evidence of interspecies transmission. Our findings suggest that these birds serve as a substantial reservoir of antigenically diverse influenza viruses, including isolates antigenically related to the current human and animal influenza viruses. This reservoir in nature may be perpetuated by a cycle involving annual infection of juvenile birds followed by transmission to the remaining susceptible birds until the next congregation during the breeding season.     

Vaccinia virus expression and sequence of an avian influenza nucleoprotein gene: potential use in diagnosis

Summary The nucleoprotein (NP) gene from avian influenza strain A/Shearwater/Aust/1/72 (H6N5) was cloned, sequenced, and expressed in vaccinia virus for the production of potent sera in immunised rabbits. The NP gene is 1565 bp and shares >95% amino acid sequence identity with other NPs of the avian subtype. The recombinant NP expressed by vaccinia virus comigrated with endogenous A/Shearwater/Aust/1/72 NP by Western blot analysis. Polyclonal rabbit sera raised against recombinant NP was evaluated in an antigen capture ELISA system as a potential diagnostic tool for the detection of avian influenza. All type A strains, comprising several HA and NA subtypes, but not type B nor other avian viruses, were detected.     

Genetic evolution of swine influenza A (H3N2) viruses in China from 1970 to 2006

Pigs are susceptible to both human and avian influenza viruses and have been proposed to be intermediate hosts, or mixing vessels, for the generation of pandemic influenza viruses through reassortment or adaptation to the mammalian host. In this study, we summarize and report for the first time the coexistence of wholly human-like H3N2 viruses, double-reassortant H3N2 viruses, and triple-reassortant H3N2 viruses in pigs in China by analyzing the eight genes of swine influenza A (H3N2) viruses found in China from 1970 to 2006. In 1970, the first wholly human-like H3N2 (Hong Kong/68-like) viruses were isolated from pigs in Taiwan, and then in the next years Victoria/75-like, Sydney/97-like, New York/99-like, and Moscow/99-like swine H3N2 viruses were regularly isolated in China. In the 1980s, two triple-reassortant viruses were isolated from pigs. Recently, the double-reassortant viruses containing genes from the human (HA and NA) and avian (PB2, PB1, PA, NP, M, and NS) lineages and the triple-reassortant viruses containing genes from the human (HA and NA), classical swine (NP), and avian (PB2, PB1, PA, M, and NS) lineages emerged in pigs in China. The coexistence of wholly human-like and reassortant viruses provides further evidence that pigs serve as intermediate hosts, or mixing vessels, and emphasizes the importance of reinforcing swine influenza virus surveillance in China.