Panorama Phylogenetic Diversity and Distribution of Type A Influenza Virus

Background Type A influenza virus is one of important pathogens of various animals, including humans, pigs, horses, marine mammals and birds. Currently, the viral type has been classified into 16 hemagglutinin and 9 neuraminidase subtypes, but the phylogenetic diversity and distribution within the viral type largely remain unclear from the whole view. Methodology/Principal Findings The panorama phylogenetic trees of influenza A viruses were calculated with representative sequences selected from approximately 23000 candidates available in GenBank using web servers in NCBI and the software MEGA 4.0. Lineages and sublineages were classified according to genetic distances, topology of the phylogenetic trees and distributions of the viruses in hosts, regions and time. Conclusions/Significance Here, two panorama phylogenetic trees of type A influenza virus covering all the 16 hemagglutinin subtypes and 9 neuraminidase subtypes, respectively, were generated. The trees provided us whole views and some novel information to recognize influenza A viruses including that some subtypes of avian influenza viruses are more complicated than Eurasian and North American lineages as we thought in the past. They also provide us a framework to generalize the history and explore the future of the viral circulation and evolution in different kinds of hosts. In addition, a simple and comprehensive nomenclature system for the dozens of lineages and sublineages identified within the viral type was proposed, which if universally accepted, will facilitate communications on the viral evolution, ecology and epidemiology.


Introduction
Influenza A virus causes frequent epidemics and occasional pandemics in various animals, including humans, pigs, horses, marine mammals and birds [1]. The viral genome comprises 8 segments which totally encode at least 10 proteins. The fourth and sixth segments encode the viral hemagglutinin (HA) and neuraminidase (NA), respectively, both of which are highly variable and diversified. Currently, 16 HA subtypes (H1,H16) and 9 NA subtypes (N1,N9) of influenza A virus have been identified [2]. Their combination further generates dozens of subtypes of influenza A virus, like H1N1, H3N2, H7N7, H9N2, etc.
Till date, the phylogenetic diversity and distribution of influenza A viruses have been widely studied within some regions, periods, hosts or viral lineages, but seldom detailed within the type or a subtype from the panorama view except for subtypes H5 and N1 influenza virus . Moreover, most of the 16 HA and 9 NA subtypes have evolved into some distinct lineages and sublineages, but it is unknown that how many lineages and sublineages exist within most of the subtypes. In addition, some of the lineages and sublineages have been designated as the North American lineage, swine human-like H1 viruses, A/chicken/Hong Kong/G9/ 97(H9N2)-like viruses, Fujian-like viruses, etc., as required by the studies on evolution, ecology and epidemiology of influenza A virus. These nomenclatures are ambiguous, miscellaneous, selflimited and sometimes even misleading. Therefore, it is of significance to identify all the lineages and sublineages within type A influenza virus and unify their nomenclature with a more rational system.
In this report, approximately 15000 HA gene sequences and 8000 NA gene of influenza A viruses available in GenBank were analyzed along with their background in order to identify all the lineages and sublineages as well as elucidate the panorama phylogenetic diversity and distribution within the whole viral type covering all the 16 HA and 9 NA subtypes. Meanwhile, a simple and comprehensive nomenclature system for the lineages and sublineages was also proposed, which if universally accepted, will facilitate international communications on the viral evolution, ecology and epidemiology.

Panorama phylogenetic diversity and distribution of type A influenza virus based on HA sequences
1264 representatives were selected from the 14328 influenza A viruses with HA sequences in GenBank. The designations of the representative viruses were partially demonstrated in Table S1 and   are available upon request. The phylogenetic tree of the representatives based on their HA sequences provided us a panorama view to the phylogenetic diversity and distribution of type A influenza virus ( Figure 1). First, most of the 16 HA subtypes could be divided into some distinct lineages which were detailed in the following sections and generalized in Table S1. Second, most of the 16 HA subtypes were distant to each other in genetics except the two pairs, H13 and H16, H7 and H15. H15 was so close to H7 that it looked like a lineage of subtype H7. Among the 16 HA subtypes, the average genetic distance between H3 and H13 was largest ( = 213.9%, ranging from 160.6% to 383.3%), and that between H13 and H16 was smallest ( = 44.3%, ranging from 40.6% to 50.6%).
Among the 16 HA subtypes, only some strains of subtypes H5 and H7 influenza viruses were found with 3 or more basic amino acid residues at the cleavage site of the HA protein which have been known a premise for high pathogenicity in birds [3]. This is consistent with the fact that only some H5 and H7 influenza viruses were highly pathogenic to birds. Among the thousands of mammalian influenza A viruses, only and all known equine H7N7 influenza viruses harbored 3 or more basic amino acid residues at the cleavage site of the HA protein.
Panorama phylogenetic diversity and distribution of type A influenza virus based on NA sequences 1154 representatives were selected from the 7836 influenza A viruses with HA sequences in GenBank. The designations of the representative viruses were partially demonstrated in Table S2 are available upon request. The phylogenetic tree of the representatives based on their NA sequences provided us another panorama view to the phylogenetic diversity and distribution of type A influenza virus ( Figure 2). First, most of the 9 subtypes could be divided into some distinct lineages which were detailed in the following sections and generalized in Table S2. Second, all the 9 subtypes are distant to each other in genetics.
The phylogenetic distribution of H1 subtype influenza viruses 215 (33 avian, 77 human, 104 swine, 1 giant anteater) representatives of the H1 influenza A viruses were selected. Their phylogenetic relationships were given in Figure 3 and consistent with the H1 part in Figure 1. Figure 3 suggested that, from a panorama view, the subtype could be divided into 3 lineages designated as h1. 1 1930,1939, 1942,1985 and 1986,2007, respectively. Lineage h1.3 mainly comprised classical swine H1 influenza viruses which circulated in the world from the 1910s until 1979 [4,9]. Thereafter, the viruses mainly circulated in America and Asia [9,10]. Sometimes, as demonstrated by Figure 3, humans or birds could be infected by the viruses of this lineage. Moreover, in 1976, more than 200 recruits in New Jersey were infected by this lineage and the abortive epidemic was feared to have pandemic   potential [6]. The lineage could be divided into two sublineages, h1.3.1 and h1.3.2, largely corresponding to the period of the 1930s,1940s and the 1970s,2000s, respectively. Only one mammalian isolate, A/swine/England/191973/ 92(H1N7) was of the NA subtype other than N1 and N2, which indicated it was an isolate resulted from genetic reassortment [11].
The phylogenetic distribution of H2 subtype influenza viruses 66 (36 avian, 1 swine and 29 human) representatives of H2 subtype influenza viruses were selected. Their phylogenetic relationships were given in Figure 4 and consistent with the H2 part in Figure 1. Figure 4 suggested that, from a panorama view, the subtype could be divided into two lineages designated as h2.1 and h2.2. Lineage h2.1 corresponded to the avian H2 influenza viruses isolated from the Western Hemisphere, while lineage h2.2 was more complicated including human and avian viruses from multiple continents. The two lineages could be further divided into some sublineages. In lineage h2.2, sublineage h2.2.6 corresponded to human H2N2 influenza viruses which circulated in the world during 1957,1968, and the rest h2.2 sublineages corresponded to the viruses isolated from the Eastern Hemisphere, except sublineages h2.2.4 surprisingly corresponded to the viruses from North America [12]. Figure 4 indicated that the HA gene of human H2N2 influenza viruses were probably originated from avian influenza viruses belonging to lineage h2.1 [13].
Panorama phylogenetic distribution of H3 subtype influenza viruses 301 (98 swine, 84 avian, 60 human, 53 equine, 3 canine, 2 environment and 1 seal) representatives of H3 subtype influenza viruses were selected. Their phylogenetic relationships were given The tree demonstrated that subtype H16 is close to subtype H13, and subtype H15 is similar to a lineage within subtype H7. The panorama diversity of each of the HA subtypes was detailed in Figure 3, Figure 9. Bootstrap values were given at relevant nodes. doi:10.1371/journal.pone.0005022.g001 in Figure 5 and consistent with the H3 part in Figure 1. Figure 5 suggested that, from a panorama view, the subtype could be divided into 3 lineages designated as h3.1, h3.2 and h3.3.
Only two representatives, one from a horse in Argentina and the other from a pig in Canada, were identified in lineage h3.3. Considering their genetic distance and distinct background, they were classified into two different sublineages, h3. 3 [20][21][22].
Because human H3 representative were selected without consideration of isolation places, the 42 human representatives actually represent much more isolates than the 30 swine representatives in h3.1.6 in Figure 5. The 30 swine representatives in h3.1.6 were similar to the human representatives isolated in the same year or earlier, which indicated that pigs might be the victims of the human influenza viruses [1].
Similar to H1 influenza viruses, Figure 5 demonstrated that interspecies transmissions within H3 influenza viruses have been identified in most of the H3 sublineages. Interestingly, some birds were found infected directly with mammalian H3N2 viruses in sublineage h3.1.6 without genetic reassortment [23,24].
Similar to human H2 influenza viruses, Figure 5 demonstrated that probably human H3 influenza viruses were originated from the avian influenza viruses circulating in the Eastern rather than Western Hemisphere.
In addition, like human H1N1 and H2N2 viruses, human H3N2 influenza viruses changed obviously with time rather than regions (data not shown).
Panorama phylogenetic distribution of H5 subtype influenza viruses 255 (1 swine, 233 avian, 19 human and 2 cats) representatives of H5 subtype influenza viruses were selected. The phylogenetic relationships of the representatives calculated with their own HA sequences were given in Figure 6 and consistent with the H5 part in Figure 1. Figure 6 suggested that, from a panorama view, the subtype could be divided into 3 lineages designated as h5.1, h5.2 and h5.3.
Only one strain, A/turkey/Ramon/73(H5N2), isolated in Mexico in 1973 was identified in lineage h5.3, while the lineages of h5.1 and h5.2 comprised many avian isolates from the Eastern and Western Hemisphere, respectively. The lineages of h5.1 and h5.2 could be further divided into several sublineages, and some sublineages could be further divided into some clades as demonstrated by the example of sublineage h5.2.3 given in Text S2.
Among lineage h5.1, the viruses within sublineage h5.1.1 were isolated in the early years, and the Mexico isolates were located in a sublineage (h5.1.6) distinct from those of USA isolates.
Among lineage h5.2, viruses within sublineage h5.2.1 were isolated in the early years, and the HPAI H5N1 widely circulating  Some H5 influenza viruses of different NA subtypes, pathogenicity or from different continents, hosts, or years could be located at the same sublineages, though they were of certain spatial and temporal features ( Figure 6).
Panorama phylogenetic distribution of H7, H10 and H15 subtypes of influenza viruses 101 H7 (87 avian, 4 human, 9 horses, 1 seal), 26 H10 (25 avian, 1 mink) and 3 H15 (all avian) representatives were selected. Their phylogenetic relationships were given in Figure 7 and consistent with their counterparts in Figure 1 except that H15 subtype was shown as an outer rather than inner branch of subtype H7.  From a panorama view, Figure 7 indicated that subtype H7 could be divided into 3 lineages, h7.1, h7.2 and h7.3. Lineage h7.3 corresponded to the equine H7N7 viruses which have not been found since the end of the 1970s [26]. Lineage h7.1 and h7.2 largely corresponded to the avian H7 influenza viruses isolated in the Western Hemisphere and the Eastern Hemisphere, respectively. Lineage h7.1 was further divided into two sublineages, h7.1.1 and h7.1.2, corresponded to the avian H7 influenza viruses isolated in North and South America, respectively. Figure 7 confirmed that the H7 influenza viruses isolated from humans and seals were all of avian sources [27][28][29][30].
Panorama phylogenetic distribution of H9 subtype influenza viruses 115 (102 avian, 8 swine, 5 human) representatives of H9 subtype influenza viruses were selected. Their phylogenetic relationships based on their HA sequences were given in    Figure 1 and previous studies on the diversity of H9 influenza viruses [31][32][33]. Figure 8 suggested that, from a panorama view, the subtype could be divided into four lineages designated as h9.1,h9.4. Lineages h9.3 and h9.4 could be further divided into some sublineages ( Figure 8) [36].

Panorama phylogenetic distribution of other HA subtypes of influenza viruses
182 representatives of other HA subtypes were selected from 635 (374 H6, 132 H4, 12 H8, 65 H11, 24 H12, 15 H13, 3 H14 and 10 H16) influenza viruses. Their phylogenetic relationships based on their HA sequences were given in Figure 9 and largely consistent with their counterparts in Figure 1.
From Figure 9, subtypes H8, H14 and H16 could not be divided into some lineages due to too few isolates available, and subtypes H4, H11 and H12 all were divided into two lineages largely corresponding to avian isolates from the Western and Eastern Hemisphere, respectively. Subtype H6 was also divided into the two lineages, but not that strictly corresponding to the two hemispheres. Subtype H13 could be divided into three lineages without obvious temporal or geographical features.
Panorama phylogenetic distribution of N1 subtype influenza viruses 441 representatives of N1 subtype influenza viruses were selected. Their phylogenetic relationships were given in Figure 10 and largely consistent with the N1 part in Figure 2. Figure 10 was consistent with a previous report on panorama of N1 influenza viruses [34]. It classified N1 influenza viruses into 3 lineages, n1.1, n1.2 and n1.3, largely corresponding to avian, human and classical swine N1 influenza virus. Each of them was further divided into several sublineages which were of certain distinct temporal, geographical or host features (Table S2). Some swine isolates formed a distinct sublineage (n1.1.7) within the avian lineage n1.1, and their HA gene sequences belonged to sublineage h1.1.3 (Figure 3). The current H5N1 HPAI viruses circulating in the Eastern Hemisphere were located in the sublineage n1.1.11.

Panorama phylogenetic distribution of N2 subtype influenza viruses
382 representatives of N2 subtype influenza viruses were selected. Their phylogenetic relationships were given in Figure 11 and largely consistent with the N2 part in Figure 2. Figure 11 classified N2 influenza viruses into 2 lineages, n2.1 and n2.2, largely corresponding to avian and mammalian N2 influenza viruses ( Figure 11A). Each of them could be divided into several sublineages which were of certain distinct temporal, geographical or host features (Table S2). Some sublineages, like n2.2.3, n2.2.4 and n2.1.4, could be further divided into some clades. A/swine/Quebec/4001/2005(H3N2) was a special isolate within N2 subtype and could be assigned as another lineage within N2 subtype ( Figure 11B).
The genetic distances between the N2 lineages and those between the N2 sublineages in the same lineage were ranging from 25 Panorama phylogenetic distribution of N3 subtype influenza viruses 65 representatives of N3 subtype influenza viruses were selected. Their phylogenetic relationships were given in Figure 12 and largely consistent with the N3 part in Figure 2. Figure 12 classified N3 influenza viruses into 2 lineages, n3.1 and n3.2. Lineage n3.1 was further divided into 3 sublineages. Sublineage n3.2 did not show any obvious geographical features. Some viruses in sublineage n3.1.1 were isolated from humans or pigs.
Panorama phylogenetic distribution of subtypes N4, N5 and N8 influenza viruses 144 representatives of subtypes N4, N5 and N8 influenza viruses were selected. Their phylogenetic relationships were given in Figure 13 and largely consistent with their counterparts in Figure 2. Figure 13 demonstrated that avian influenza viruses of these three subtypes all could be classified into 2 lineages corresponding to the Western Hemisphere (n4.1, n5.1 and n8.1) and Eastern Hemisphere (n4.2, n5.2 and n8.2), respectively, with some exceptions in sublineage n8.1. The equine N8 influenza viruses formed a separate lineage within subtype N8.

Discussion
The topologies of the phylogenetic trees described here were all consistent with, though much more covering than, previous reports . During the analysis of the sequences, we found that the topology of phylogenetic trees changed little if calculated with different software packages like MEGA, PHYLIP or PAUP, or calculated with different methods like neighboring-joining or maximum parsimony (data not shown). But it could change greatly using different parameters. In this study, we set the nucleotide sequences changing in Kimura-2-Parameter gamma model because we found that transitional substitution rates were much higher than transversional ones in the viral evolution, and substitution rates were significantly different among sites. In addition, it is important to treat gaps by pairwise-deletion rather than complete-deletion, i.e. ignoring only those gaps that are involved in the comparison of a pair of sequences because random insertions or deletions were frequent in the evolution of the two viral genes especially among different subtypes.
During the preliminary analysis of the sequences, we found that the trees might change greatly if some short sequences were involved. Therefore, in this study, sequences shorter than a limitation were excluded.
It is possible that the panorama views reported here reflected only a part of the reality. First, few isolates have been reported from South America and Africa, and influenza viruses of marine mammals also remain largely unknown. Second, it is possible that a few sequences reported to GenBank were with some errors which could distort the phylogenetic ''maps'' described here. Nevertheless, the panorama maps help us to recognize influenza A viruses from panorama views, and provided a framework for to generalize the history and explore the future of the viral circulation in multiple kinds of hosts. For example, the pandemic of human H3N2 virus in 1968 [19], the severe outbreak of equine influenza with an avian-like H3N8 virus in China in 1989 [35], and the crisis of H5N1 HPAI in Hong Kong in 1997 [36], all could be marked in the maps. As for the future, it is easier with the guide of the panorama maps to identify whether an isolate is special or odd in epidemiology (like A/equine/Argentina/1/01(H3N8) in lineage h3.3.1 in Figure 5), and whether an isolate originated from genetic reassortment, like the aforementioned A/swine/England/ 191973/92 (H1N7) [11].
The panorama maps gave us some new information to recognize influenza A virus. First, they demonstrated that H13 and H16 were very close in genetics, and so did subtype H7 and H15. The maps also indicated that some subtypes like H3, H5, H7 and H9 might have evolved into some subtypes during the evolution in the past decades, which, however, should be confirmed by serological tests. Second, avian influenza viruses of each subtype were usually classified into Eurasian and North American lineages in the past presumably owing to confinement of birds to the distinct flyways of each hemisphere [37][38][39], and here the two lineages were confirmed within H1, H3, H5, H7, H8, H10, N4, N5, N6 and N8 subtypes, but their geographical distribution should be enlarged to the Eastern (Asia, Oceania, Europe and Africa) and Western Hemisphere (North and South America), respectively. Moreover, the avian influenza viruses within subtypes H2, H4, H6, H9, H11, H13, N1, N2, N3, N7 and N9 were more complicated than these two lineages. For example, avian H9 influenza viruses should be divided into 4 lineages, two for each hemisphere, and some avian viruses isolated from the Western Hemisphere was located in one sublineage (h9.3.2) of the lineage h9.3 corresponding to the Eastern Hemisphere.
The panorama maps demonstrated that swine H1 and H3 viruses were of high diversity, and swine infections with avian or human H1 and H3 influenza viruses as well as human or avian infections with swine H1 and H3 viruses were rather frequent. Therefore, the maps offered us a new dimension to recognize that pigs may play a important role in the ecology and epidemiology of H1 and H3 influenza viruses.
The panorama maps also demonstrated that unlike swine and avian influenza viruses, human influenza viruses demonstrated obvious temporal but little geographical difference, and they have diverged into few clades during the past decades. These suggested that human viruses update rapidly and they were maintained almost exclusively by humans themselves. Similar features were also found in equine influenza viruses. The features indicated that the circulation of human and equine influenza A viruses was vulnerable, which was supported by the stop of the circulation of human H2N2 and equine H7N7 influenza viruses [1].
The phylogenetic distribution reported here suggests that, from a whole view, there is a little possibility for an avian influenza virus to infect humans or other mammals, but little possibility for a mammalian influenza virus to infect birds, though some bird infections with swine H1N1 viruses in sublineage h1.3.2 and H3N2 viruses in sublineage h3.1.6 have been found. Therefore, the panorama analysis supported that influenza viruses lost their adaptability in birds while they have become adapted to mammalian hosts [1].
The panorama phylogenetic trees are useful for detection of the viruses. For example, the primers of the current China national standard RT-PCR for the detection of H7 subtype avian influenza viruses, 5-AATGCACARGGAGGAGGAACT-3 (upper) and 5-TGAYGCCCCGAAGCTAAACCA-3 (down), are specific to lineage h7.2 rather than h7.3, while the later corresponds to the avian H7 influenza viruses isolated in the Eastern Hemisphere and should be more risky to China according to their phylogenetic distribution. Therefore, the primers should be revised to those corresponding to lineage h7.3 or both h7.2 and h7.3. For another example, a few pairs of primers may be required to detect swine H1 or H3 influenza viruses using RT-PCR as they are of high diversity demonstrated by the panorama views. In addition, the representative sequences selected by this report could give a window to identify the specific and conserved regions in the viral HA and NA sequences for the primer design.
Classification and designation of the lineages and sublineages within influenza A virus were essential for the studies of the viral evolution, ecology and epidemiology [1,. In this report, 60 lineages and 83 sublineages within influenza A virus were identified, and all of them were supported by the topology of the phylogenetic trees and with high bootstrap values (.70) at relevant nodes except lineage n1. 2 [40].
In this report, we tentatively proposed a kind of nomenclature for the lineages and sublineages within influenza A virus. It is informative and specific as it begins with the subtype information. Because only two hierarchies are involved in the nomenclature for simplicity, some sublineages were probably evolved from other sublineages, like sublineage h5.2.3 was possibly evolved from sublineage h5.2.2 which, in turn, was possibly evolved from sublineage h5.2.1 at least in terms of the HA gene ( Figure 6). Meanwhile, some sublineage could be further divided into some distinct clades as shown by the example given in Text S2.
The nomenclature reported here covers nearly all known influenza A viruses, and it is easy to be expanded to meet the future evolution of the viruses. For example, a new lineage, n2.3, could be assigned to the special isolate in Figure 11B, and some new sublineages could be assigned to the viruses, A/duck/NZL/ 38/1984 and A/swine/Chonburi/05CB2/2005 in Figure 5. Moreover, the WHO/FAO/OIE classification and designations of the clades and subclades within the sublineage h5.2.3 (Text S2) also provided a kind of expansion of the nomenclature of the lineages and sublineages reported here.
If this nomenclature is widely accepted, it should facilitate international communication on the evolution, ecology and epidemiology of influenza A viruses through the unification of previous miscellaneous nomenclatures which were not only ambiguous or misleading in some cases, but also covered only a part of the diversity of influenza A viruses. For example, the previous designation ''swine human-like H1 viruses'' was ambiguous as the swine viruses within the human H1 lineage belonged to multiple sublineages (Figure 3), and the previous designation ''Eurasian lineage of H2 influenza virus'' actually comprised viruses from Australia and Africa and even some from North America (Figure 4). Moreover, a substantial part of subtypes H3, H5, H9, N3, etc., have not or could not be covered by previous nomenclatures.

Selection and primary analysis of representative sequences
All the around 15000 HA and 8000 NA sequences of influenza A viruses available in GenBank till July 11, 2008 were analyzed firstly through the web servers of Influenza Virus Resource in NCBI [41], to exclude the sequences of the same viruses, the viruses with unclear background, or manipulated materials and the sequences shorter than 500 bp or with sequencing errors like the ones with GenBank accession numbers AX822028, AX822031, AX822045. The rest sequences were then utilized for further analysis (assortment, alignment and tree-building) through the web servers of Influenza Virus Resource to elucidate their host and subtype distributions and identify the number of basic amino acid residues at the cleavage site in the HA sequences.
The representative sequences were selected by randomly choosing one from those which were similar to each other (approximately genetic distances ,2.1%), isolated from the similar host (avian or a species of mammals) in the same year and in the same country. Since human H1N1, H2N2 and H3N2 subtypes viruses did not shown obvious geographical bias through primary analysis and they were of large numbers, their representatives were selected by randomly choosing one from those which were similar to each other and isolated in the same year no matter where they were isolated.

Calculation of genetic distances
Genetic distances among the representative sequences were calculated using the model of Kimura-2-Parameter using the software MEGA 4.0 [42]. The substitution rates among sites were set in Gamma distribution. The Gamma parameter was set as 1.0. Gaps were treated by pairwise-deletion, i.e. ignoring only those gaps that are involved in the comparison of a pair of sequences. The genetic distances between lineages and those between sublineages were calculated through the comparison of the selected representative sequences.

Phylogenetic analysis
Representative sequences were aligned using the software of Clustal X [43], and their phylogenetic relationship was analyzed using the software MEGA 4.0 with neighboring-joining method and the same parameters for the genetic distance calculation [42]. with neighboring-joining method [42] using the same parameters for the genetic distance calculation. Bootstrap values were calculated out of 1000 replicates. All the phylogenetic trees were calculated by two separate groups to avoid manual errors.

Classification of lineages and sublineages
Lineages and sublineages were classified according to genetic distances and topology of the phylogenetic trees. Distributions of the viruses in hosts, regions and time were also considered in the classification. Lineage designation began with the subtype name which was followed by a point and a number (e.g. n2.1), and sublineage designation began with its lineage designation which was followed by a point and a number (e.g. n2.1.1). The numbers within the designations of the lineages and sublineages, if possible, were in the order of avian, human, swine, equine followed by others in terms of hosts, and in the order of the Western Hemisphere followed by Eastern Hemisphere in geography, and in the order from the past to nowadays in isolation time.