Detection and characterization of new influenza B virus variants in 2002

One-hundred five influenza B-positive specimens obtained from southeast Asia in 2002 were categorized on the basis of DNA sequencing of HA1 gene as well as real-time PCR analysis of the NA gene. Phylogenetic analysis of the HA1 gene sequences showed that the majority of the viruses (96.2%) belonged to the B/Victoria/2/87 lineage, while a smaller percentage of the viruses (3.8%) belonged to the B/Yamagata/16/88 lineage. The B/Yamagata/16/88 viruses displayed significant antigenic drift in the deduced amino acid sequences of the HA1 protein, and the B/Victoria/2/87-like viruses consisted of B/Hong Kong/1351/02-like (72.3%) and B/Hong Kong/330/01-like (27.7%) viruses. The B/Hong Kong/1351/02-like viruses were reassortants with the HA gene belonging to the B/Victoria/2/87 lineage and the NA gene belonging to the B/Yamagata/16/88 lineage, whereas both the HA and NA genes of B/Hong Kong/330/01 virus belonged to the B/Victoria/2/87 lineage. In this study, however, all the B/Hong Kong/330/01-like isolates exhibited the B/Yamagata/16/88-like NA gene, which likely resulted from reassortment of B/Hong Kong/330/01 and B/Hong Kong/1351/02 viruses during coinfection. Additional molecular characterization of the six internal genes showed that the M, NS, PA, and PB2 genes of the new variants were B/Hong Kong/1351/02 in origin, whereas the NP and PA genes retained the B/Hong Kong/330/01 origin. Interestingly, these new variants all appeared late in the year 2002. These results support the notion that influenza B viruses continued to evolve through antigenic drift and shift.     

Changes in the hemagglutinins and neuraminidases of human influenza B viruses isolated in Italy during the 2001-02, 2002-03, and 2003-04 seasons

Throughout most of the last decade, B/Yamagata/16/88-lineage influenza viruses were predominant among the B isolates circulating worldwide, whereas B/Victoria/2/87-lineage viruses were isolated infrequently and restricted geographically to eastern Asia. During the 2001-02 influenza season, B/Victoria/2/87-lineage viruses re-emerged in North America and Europe and spread worldwide. Virological surveillance in Italy during that season showed wide circulation of influenza B viruses, of which most were antigenically related to the B/Sichuan/379/99 (Yamagata-lineage) vaccine strain, together with a smaller number of B viruses antigenically similar to B/HongKong/330/01, a recent B/Victoria/2/87-lineage antigenic variant. In the subsequent 2002-03 epidemic season, B viruses with a Victoria-lineage hemagglutinin (HA), more closely related to that of B/Shandong/7/97, were isolated exclusively. Similar strains have continued to predominate among the few B viruses isolated in Italy during last season (2003-04), although most influenza B viruses, isolated sporadically elsewhere in Europe, again belong to the Yamagata-lineage. In the present study, phylogenetic analyses of the HA and neuraminidase (NA) genes of representative B strains, isolated throughout Italy during 2001-04, showed that during the first influenza season the NA genes, as well as the HA genes, separated into the two distinct clades, the Yamagata- and Victoria-lineages, and showed no evidence of genetic reassortment. On the contrary, all the B viruses isolated in the 2002-03 and most of those isolated in the 2003-04 epidemic season were "Victoria HA-Yamagata NA" reassortants similar to those isolated in other parts of the world, showing that these reassortants became established in the human population. The frequency of reassortment between HA and NA of distinct lineages and sublineages highlights again the importance of detailed molecular analyses of both surface glycoproteins in understanding the evolution of influenza B viruses.     

Increasing appearance of reassortant influenza B virus in Taiwan from 2002 to 2005

Genetic and antigenic analyses of influenza B virus field strains isolated in Taiwan from 1998 to 2005 were performed. To investigate the molecular evolution of influenza B viruses, sequence analysis of the hemagglutinin (HA1 subunit) and neuraminidase genes was performed. All influenza B viruses isolated between 1998 and 2000 belonged to the B/Yamagata/16/88 lineage. The B/Victoria/2/87 lineage, which was cocirculating with the Yamagata lineage, was identified in Taiwan in March 2001. Concurrently, there was an increasing prevalence of this lineage in many parts of the world, including North America and Europe, during the 2001-2002 season. Since 2002, genetic reassortants of influenza B virus with the Victoria lineage of hemagglutinin and the Yamagata lineage of neuraminidase have been found at a rate of 46%. Therefore, in 2002, at least three sublineages of influenza B virus strains, the B/Shanghai/361/2002-like strain (Yamagata lineage), the B/Hong Kong/330/01-like strain (Victoria lineage), and the B/Hong Kong/1351/02-like strain (B reassortant lineage), were identified in Taiwan. The results showed that genetically distinct lineages can cocirculate in the population and that the reassortment among these strains plays a role in generating the genetic diversity of influenza B viruses. Interestingly, from January to April 2005, B reassortant viruses became dominant (73%) in Taiwan, which indicated that a mismatch had occurred between the influenza B vaccine strain recommended for the 2004-2005 season in the Northern hemisphere by the World Health Organization and the epidemic strain.     

Cocirculation and evolution of two lineages of influenza B viruses in europe and Israel in the 2001-2002 season

Forty-nine influenza B virus isolates collected in Belgium, Finland, Spain, and Israel during the 2001-2002 winter season were categorized into either of two lineages, B/Yamagata/16/88 or B/Victoria/2/87, based on the phylogenetic studies of HA1 sequences. The data trace the geographic spread of B/Victoria/2/87-like viruses and support the emergence of B/Hong Kong/1351/02-like viruses, possibly due to selective advantages of reassortment.     

Influenza viruses which preconditioned the epidemic rise in Russia in 2002-2003. A resumed circulation of influenza viruses similar to V/Victoria/2/87

According to research, the epidemic rise of influenza was preconditioned, during 2002-2003, in Russia by the circulation of influenza A(H1N1), A(H3N2) and B viruses. The Center of Influenza Ecology and Epidemiology undertook a study of 178 epidemic strains: 41 strains A(H1N1), 116 strains A(H3N2) and 21 strains of influenza B were among them. All strains were isolated in the MDCK cell culture. A simultaneous isolation in embryonated eggs as well as changing of the isolation system from MDCK to embryonated eggs were found to be effective only for influenza A(H1N1) viruses. According to the antigenic analysis, all A(H1N1) viruses were variants of the etalon A/New Caledonia/20/99. The A(H3N2) viral strains' population was heterogeneous by its antigenic properties: among its isolates, there were variants similar to the etalons of A/Moscow/10/99 and of A/Panama/200/99 as well as strains, which weakly reacted with sera of both above etalons; possibly the latter were close to the etalon of A/Fujian/411/02. All epidemic strains of influenza B virus belonged, according to the antigenic properties of hemagglutinin, to the virus group of B/Victoria/2/87-like and were antigenic variants of the etalon of B/Hong Kong/22/01. This confirmed that influenza B viruses with the antigenic hemagglutinin structure of the virus group of B/Victoria/2/87-like, which were not present in Russia for more than 10 years, re-entered the active circulation. An analysis of antigenic properties of neuraminidases (NA) of the mentioned epidemic strains showed their different degrees of relationship with the NA etalons of both evolutionary groups, i.e. B/Victoria/2/87 and B/Yamagata/16/88-like. A study of paired sera obtained from patients showed a growth of antibodies to the etalons of influenza A(H1N1), A(H3N2) and B viruses of the season in question, which confirmed the virology data.     

Recent changes among human influenza viruses

Recurrent epidemics of influenza are due to the frequent emergence of antigenic variants. With co-circulation of two influenza A subtypes and two antigenically distinct lineages of B viruses, genetic reassortment also has an important role in antigenic drift, as illustrated by recent changes in both A and B viruses. The H1N2 subtype viruses, which emerged during 2001, possessed a H1 HA similar to those of contemporary A/New Caledonia/20/99 (H1N1)-like viruses and seven genes closely related to those of recent H3N2 viruses, and did not represent a significant increase in the antigenic diversity of circulating viruses. The re-emergence of B/Victoria/2/87-lineage viruses, previously prevalent during the 1980s, in 2000 has been followed by the predominant circulation of reassortant B viruses possessing a B/Victoria-lineage HA and a B/Yamagata-lineage NA similar in sequence to those of recent B/Sichuan/379/99-like viruses. These events emphasize not only the lack of divergence in the complementary functional characteristics of the HA and NA of divergent influenza B lineages, but also the apparent convergence in compatibility between the H1 and N2 components of the two influenza A subtypes.     

Reassortment and evolution of current human influenza A and B viruses

During the 2001-2002 influenza season, human influenza A (H1N2) reassortant viruses were detected globally. The hemagglutinin (HA) of these H1N2 viruses was similar to that of the A/New Caledonia/20/99 (H1N1) vaccine strain both antigenically and genetically, while their neuraminidase (NA) was antigenically and genetically related to that of recent human influenza H3N2 reference viruses such as A/Moscow/10/99. All six internal genes of the H1N2 reassortants originated from an H3N2 virus. After being detected only in eastern Asia during the past 10 years, Influenza B/Victoria/2/87 lineage viruses reappeared in many countries outside of Asia in 2001. Additionally, reassortant influenza B viruses possessing an HA similar to that of B/Shandong/7/97, a recent B/Victoria/2/87 lineage reference strain, and an NA closely related to that of B/Sichuan/379/99, a recent B/Yamagata/16/88 lineage reference strain, were isolated globally and became the predominant influenza B epidemic strain. The current influenza vaccine is expected to provide good protection against H1N2 viruses because it contains A/New Caledonia/20/99 (H1N1) and A/Panama/2007/99 (H3N2) like viruses whose H1 HA or N2 NA are antigenically similar to those of recent circulating H1N2 viruses. On the other hand, widespread circulation of influenza B Victoria lineage viruses required inclusion of a strain from this lineage in influenza vaccines for the 2002-2003 season.     

Evolution of Surface and Nonstructural-1 Genes of Influenza B Viruses Isolated in the Province of Québec,Canada, during the 1998–2001 Period

After 2 minor winter seasons, influenza B viruses were predominantly isolated in the Province of Quebec, Canada, during the 2000–2001 season representing 74% of laboratory-confirmed influenza viruses. We performed an antigenic study of the hemagglutinin (HA) protein and a molecular characterization of the HA1 region, nonstructural-1 (NS1) and neuraminidase (NA)/NB genes of 20 influenza B strains isolated in the Province of Quebec during the 1998–2001 period. Our isolates were compared to recent vaccine strains (B/Harbin/7/94 in 1998–1999, B/Yamanashi/166/98 in 1999–2000 and 2000–2001, and B/Sichuan/379/99 in 2001–2002). The hemagglutination inhibition (HI) test revealed that all isolates were different from B/Harbin/7/94 and were more related to the 2 other vaccine strains although precise identification was often impossible. Molecular analysis of the HA1 gene revealed that both B/Yamanashi/166/98-like and B/Sichuan/379/99-like isolates co-circulated during the 1998–1999 season whereas isolates from the 2 subsequent years were more related to B/Sichuan/379/99. Most isolates (8/9) of the 2000–2001 season contained a N126D substitution recently associated with altered antigenicity in recent influenza B/Yamagata/16/88-related viruses. Although the HA1 and NS1 protein sequences of viruses isolated during the 1998–1999 season were clearly different from those of the respective vaccine strain (B/Harbin/7/94), the NA protein sequence of those isolates was slightly more related to B/Harbin/7/94 than B/Yamanashi/166/98 suggesting distinct patterns of evolution for these genes. This study confirms the importance of a detailed molecular analysis for understanding the evolution of influenza B viruses.     

Molecular analysis of isolates from influenza B outbreaks in the U.S. and Nepal, 2005

Summary. Currently circulating influenza B viruses can be divided into two antigenically and genetically distinct lineages referred to by their respective prototype strains, B/Yamagata/16/88 and B/Victoria/2/87, based on amino acid differences in the hemagglutinin surface glycoprotein. During May and July 2005, clinical specimens from two early season influenza B outbreaks in Arizona and southeastern Nepal were subjected to antigenic (hemagglutinin inhibition) and nucleotide sequence analysis of hemagglutinin (HA1), neuraminidase (NA), and NB genes. All isolates exhibited little reactivity with the B/Shanghai/361/2002 (B/Yamagata-like) vaccine strain and significantly reduced reactivity with the previous 2003/04 B/Hong Kong/330/2001 (B/Victoria-like) vaccine strain. The majority of isolates were antigenically similar to B/Hawaii/33/2004, a B/Victoria-like reference strain. Sequence analysis indicated that 33 of 34 isolates contained B/Victoria-like HA and B/Yamagata-like NA and NB proteins. Thus, these outbreak isolates are both antigenically and genetically distinct from the current Northern Hemisphere vaccine virus strain as well as the previous 2003–04 B/Hong Kong/330/2001 (B/Victoria lineage) vaccine virus strain but are genetically similar to B/Malaysia/2506/2004, the vaccine strain proposed for the coming seasons in the Northern and Southern Hemispheres. Since these influenza B outbreaks occurred in two very distant geographical locations, these viruses may continue to circulate during the 2006 season, underscoring the importance of rapid molecular monitoring of HA, NA and NB for drift and reassortment.     

The properties of the epidemic influenza viruses A and B strains circulating in Russia in the 2004-2005 epidemic season

The epidemic upsurge of influenza morbidity in Russia in 2004-2005 was caused by the active circulation of influenza A(H3N2) and B viruses. A hundred and sixty-six epidemic strains were studied. All the strains were isolated in the MCK cell culture. Influenza A(H3N2) viruses (n=77) were antigenic variants of the reference A/Fujian/411/ 2002 and A/California/7/2004 strains. Three influenza A(H1N1) viral strains that were antigenic variants of A/New Caledonia/20/99 strains were isolated in sporadic cases. Influenza B virus strains (n=83) were antigenic variants of the reference B/Shanghai/361/02--lineage B/Yamagata/l6/88. In addition, 3 antigenic variants of B/Hong Kong/ 330/2002 (lineage B/Victoria/2/87) strains were isolated. Nine (20%) strains resistant to rimantadine at a concentration of 5 microg/ml were identified. Chromatographic analysis of B/Shanghai/361/02 and BIHong Kong/330/01 viral protein M1 trypsin hydrolysates revealed differences in the profiles of chromatograms of influenza A virus proteins M1. Examination of 121 pair sera from patients revealed an increase in antibodies to influenza A(H3N2) viruses in 10-21% of cases and to influenza B viruses in 20-36% of cases.     

Cocirculation of two distinct evolutionary lineages of influenza type B virus since 1983

During 1988-1989 two highly distinct antigenic variants of influenza type B were recognized in hemagglutination-inhibition tests with postinfection ferret serum. These viruses were antigenically related to either B/Victoria/2/87, the most recent reference strain, or B/Yamagata/16/88, a variant that was isolated in Japan in May 1988. All influenza B viruses isolated in the United States during an epidemic in the winter of 1988-1989 were antigenically related to B/Victoria/2/87. However, in several countries in Asia, both B/Victoria/2/87-like viruses and B/Yamagata/16/88-like viruses were isolated. Sequence analysis of the hemagglutinin (HA) genes of several influenza B isolates from 1987 to 1988 indicated that the HA1 domains of the B/Yamagata/16/88-like viruses and B/VI/87-like viruses isolated in 1988 differed by 27 amino acids. Evolutionary relationships based on this sequence data indicated that the B/Yamagata/16/88-like viruses were more closely related to epidemic viruses from 1983 (B/USSR/100/83-like viruses) than to more recent reference strains such as B/Victoria/2/87. All other Asian strains, as well as selected isolates from the United States in 1988, were confirmed by sequence analysis as being genetically related to B/Victoria/2/87. These data provide clear evidence that two parallel evolutionary pathways of influenza type B have existed since at least 1983 and that viruses from each of the separate lineages were isolated from cases of influenza B in 1988. This finding is similar to earlier observations for type A H1N1 and H3N2 influenza viruses.     

Genetic diversity of influenza B virus: the frequent reassortment and cocirculation of the genetically distinct reassortant viruses in a community

To characterize the genetic diversity of influenza B viruses isolated during one influenza season, the antigenic and genetic relationships among 20 strains of influenza B virus isolated in February and March 2001 at one pediatric clinic in Yamagata City, Japan, were investigated. The HA gene and seven other gene segments were phylogenetically divided into three distinct sublineages (Harbin/7/94-, Tokyo/6/98-, and Shiga/T30/98-related lineage) of the Yamagata/16/88-like lineage. The NS genes of the viruses belonging to the Harbin/7/94-related lineage have additional three nucleotides at positions 439-447, and were phylogenetically distinguishable from those of the currently circulating Yamagata/16/88- and Victoria/2/87-like lineages, but were closely related to that of the Yamagata/16/88-like lineage isolated before 1994. Moreover, four strains of influenza B virus isolated in the same community between 2002 and 2003 were further examined. Phylogenetic analysis revealed that a virus of Victoria/2/87-like lineage isolated in 2003 had acquired the NA, NS, M, and PA gene segments from a Shiga/T30/98-like virus, and two strains of Harbin/7/94-related lineage had acquired the various gene segments from Shiga/T30/98-like virus through a reassortment event. These results indicate that genetically distinct multiple viruses can combine to cause an influenza B epidemic in a community and that the frequent reassortment among these viruses plays a role in generating the genetic diversity of influenza B viruses.     

Internal epidemic influenza virus proteins: isolation and investigation

The internal influenza virus proteins M1 and RNP free from surface protein impurities were isolated from subviral particles (virions free from HA and NA ectomenes). The spikeless particles had no propensity to aggregate in the solution at pH 5.0 as compared with native viruses. The subviral particles of B/Hong Kong/330/01 influenza virus, which belonged to B/Victoria/2/87-lineage, were obtained by proteolytic treatment with the enzyme bromelain under the same conditions as in cases of influenza B viruses of B/Jamagata/16/88 lineage. A chromatographic analysis of the tryptic hydrolyzates obtained for matrix (M1) proteins of A(H1N1) and A(H3N2) influenza viruses revealed differences that were greatest between the protein M1 molecules isolated from influenza viruses of different subtypes of hemagglutinine. These findings suggest there are variations in the structure of this conservative internal viral protein M1 during evolution.     

The reappearance of Victoria lineage influenza B virus in Brazil, antigenic and molecular analysis.

BACKGROUND: In contrast to influenza A, minor influenza B viruses can co-circulate with the dominant strain during an epidemic allowing the re-emergence of old strains and reassortment between those different strains. The 2001-2002 influenza season in the northern hemisphere was distinguished by the re-emergence of the Victoria-lineage viruses, which replaced the Yamagata-lineage, after being restricted to East Asia throughout the 1990s. OBJECTIVES: To describe the antigenic and genetic characteristics of influenza B viruses detected in South and South East Brazil and determine their lineages. STUDY DESIGN: Influenza samples collected during epidemics between 1999 and 2002 were analyzed by indirect immunofluorescence assay (IFA). Positive results were confirmed through multiplex PCR and isolation in cell culture. Isolated viruses were antigenically characterized by hemagglutination inhibition. Fourteen hemagglutinin (HA) gene sequences obtained in this work were used for phylogenetic analysis. RESULTS: Brazilian isolates from 2002 were associated with the Victoria-lineage, diverging from the vaccine used throughout that influenza season in Brazil. CONCLUSIONS: These results indicate the reappearance of Sichuan/7/97-like samples in South and South East Brazilian Regions simultaneously. They indicate the need for neuraminidase gene evaluation and demonstrate the importance of influenza laboratory surveillance to establish which strains should be included in the influenza vaccine.     

Evolutionary characteristics of influenza B virus since its first isolation in 1940: dynamic circulation of deletion and insertion mechanism

Summary.  New antigenic variants of B/Yamagata/16/88-like lineage which appeared in the season of 1997 as a minor strain tended to predominate in the following season. Also, we could observe for the first time, three peaks of activity caused by H3N2 virus and two variants of B influenza virus. Antigenic and phylogenetic analyses revealed that B/Victoria/2/87-like variants appeared again in Japan in 1997 after a nine-year absence. Influenza B viruses evolved into three major lineages, including the earliest strain (I), B/Yamagata/16/88-like variants (II), which comprised of three sublineages (II-(i), II-(ii), II-(iii)), and B/Victoria/2/87-like variants (III). Evolution of influenza B virus hemagglutinin was apparently distinguishable from that of influenza A virus, showing a systematic mechanism of nucleotide deletion and insertion. This phenomenon was observed to be closely related to evolutionary pathways of I, II-(i), II-(ii), II-(iii) and III lineages. It was noteworthy to reveal that the nucleotide deletion and insertion mechanism of influenza B virus completed one cycle over a fifty-year period, and that a three nucleotide deletion was again observed in 1997 strains belonging to lineage II-( iii). It was evident that amino acid substitutions accompanying nucleotide insertions were highly conserved. Received December 4, 1997 Accepted March 10, 1998     

Antigenic and genetic analyses of influenza B viruses\\ isolated in Lusaka, Zambia in 1999

Summary.  Previous studies of the hemagglutinin (HA) genes of various influenza B virus isolates demonstrated the existence of two antigenically distinct virus lineages represented by B/Victoria/2/87 and B/Yamagata/16/88, respectively. Here, we investigated the antigenic and genetic characteristics of influenza B viruses isolated from children living in Lusaka, Zambia between January and May 1999. Antigenic analysis with chicken antiviral sera showed that all the Zambian isolates had the HA protein belonging to B/Yamagata/16/88-related lineage. Furthermore, phylogenetic analyses of the eight RNA segments performed by using the total or partial nucleotide sequences of the two representative Zambian strains (B/Lusaka/270/99 and B/Lusaka/432/99) as well as the previously reported sequences suggested that the Zambian viruses are closely related to the recently circulating reassortants represented by B/Shiga/T30/98 and B/Yamanashi/166/98 which acquired the genes coding for three polymerase proteins (PB2, PB1, and PA), HA, nucleoprotein, and matrix protein from a B/Yamagata/16/88-like parent and the gene encoding nonstructural proteins (NS1 and NS2) from a B/Guandong/8/93-like parent. Accepted June 15, 2001 Received April 17, 2001     

Reactogenicity and immunogenicity of a surface-antigen-adsorbed influenza virus vaccine in children.

An influenza virus vaccine containing the purified surface haemagglutinin and neuraminidase antigens of A/Victoria/75 and B/Hong Kong/73 viruses adsorbed to an aluminium hydroxide gel was assessed for reactogenicity and immunogenicity in children aged 4 to 11 years, since there is no influenza virus vaccine available for this age group. Significant serum haemagglutination-inhibiting antibody responses to the A/Victoria/75 and B/Hong Kong/73 haemagglutinin antigens present in the vaccine were observed in 47% and 35%, respectively, of the children vaccinated, with a single dose. The vaccine induced no significant local or systemic reactions.     

Reassortants in recent human influenza A and B isolates from South East Asia and Oceania

From 2000 to 2002, human influenza A and B viruses that were genetic reassortants of contemporary circulating human strains, were isolated in South East Asia and Oceania. Similar to reports from other regions, A(H1N2) isolates were found to be reassortants of circulating A(H3N2) viruses that had acquired only the haemagglutinin gene of an A(H1N1) virus. Some of these reassortants from Thailand and Singapore predate those previously recorded during the winter of 2001-2002 in Europe and the Middle East and may be precursors of these viruses. The B reassortants had a haemagglutinin similar to an earlier B strain, B/Shangdong/7/97 (B/Victoria/2/87-lineage) and a neuraminidase similar to the recently circulating B/Sichuan/379/99 virus (B/Yamagata/16/88-lineage). Despite the early occurrences of A(H1N2) reassortants and the extensive circulation of A(H1) viruses in South East Asia and Oceania during 2000-2001, these reassortant influenza A viruses have to date not been prominent unlike Europe and the Middle East where they were common in the 2001-2002 winter. In contrast the reassortant B viruses, which first emerged in this region in early 2002, rapidly became the predominant strains isolated from patients with influenza B in South East Asia and Oceania.     

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.     

Genetic analysis of an influenza B virus isolated from a patient with encephalopathy in Japan

An influenza B virus, B/Saga/S172/99 (SAG99), was isolated from the nasopharynx of a patient with encephalopathy/encephalitis in Japan in 1999. To clarify the molecular characteristics of this virus, detailed analysis of the gene segments coding for the hemagglutinin (HA), neuraminidase (NA), nucleoprotein (NP), matrix protein (M) and non-structural protein (NS) was undertaken. All five genes of SAG99 showed high nucleotide and predicted amino acid similarities with those of recent non-encephalopathic strains isolated in the same epidemic season. Subsequent phylogenetic analysis revealed that all five gene segments of SAG99 analyzed in the present study were most similar to those of the recent Yamagata/16/88-like viruses. The hemagglutinin and neuraminidase proteins of SAG99 were each distinguished from those of recent epidemic strains by one characteristic amino acid substitution. These substitutions were not found in the previously reported encephalopathy/encephalitis-derived influenza B viruses, and we could not find any common characteristic amino acid changes in SAG99 and these viruses. Similarly, among the internal proteins studied, only the M2 protein of SAG99 was found to contain a single novel amino acid change when compared with other recent isolates. Thus, it was apparent that SAG99 contained very few amino acid differences when compared with other epidemic viruses. The association of recent B/Yamagata/16/88-like viruses with encephalitis/encephalopathy observed in the present study and previously suggest that these viruses may have a higher potential for causing neurological complications in certain individuals.