Discussion
Highly pathogenic AIVs have been affecting the world’s poultry industry
and also brought enormous challenges to the public health security since
they accidentally moved from animals to human, causing a first outbreak
in Guangdong, China, in 1996(Shortridge et al., 1998; Webster &
Govorkova, 2014; Xu et al., 1999). Fortunately, there is no evidence of
human-to-human transmission of the avian influenza virus. The H5 avian
influenza virus has evolved rapidly from Clade 0 to the present Clade
2.3.4.4 and Clade 2.3.2.1(Group, 2012), and frequent reassortments of
AIVs have been the leading cause of several epidemics(Bi et al., 2016;
Karo-karo et al., 2019; Qi et al., 2018; Shi et al., 2018). The highly
pathogenic avian influenza virus prevalence in China is an
extraordinarily complicated problem to solve. We focused on the
prevalence, gene differentiation, and reassortment of H5N6 in China,
based on the available data for 2016−2019. We gathered evidence on a new
reassortment virus complex diversity produced by frequent reassortment
of different subtypes AIVs. Cases of both poultry and human infections
are largely related to the complexity of the reassortment.
The rate of evolution of avian influenza also deserves close attention.
The rate of evolution of H5N6 HA and NA genes is 7.3262E-03 and
6.9073E-03, respectively. And the rate of evolution of H9N2 HA and NA is
2.19E-03-2.83E-03, and is
3.3E-03-3.7E-03, respectively(Jin et al., 2020). H5N6 compared to H9N2
has a faster rate of evolution in HA and NA genes.This has a lot to do
with the clade of HA and NA genes rapid differentiation is closely
linked. And internal gene is relatively complicated, due to the
restructuring of the LAIVs, especially the part of basic internal genes
from H6N6, H6N2, H9N2, most notably the two H6N6 viruses from which the
immune pressure is relatively small. According to the study of Bi et
al(Bi et al., 2016), the PB1 gene of H5N6 is basically derived from
H5N1, and PB2 is partly derived from H6N6, which may be the reason why
our study showed that PB1 has significant gene differentiation compared
with other genes.
In this study, a total of 16 gene sequences from humans [Figure 6
A], including eight strains isolated during 2016 (six isolates from
January to April and two from November and December), two isolated since
November, seven of eight strains were new reassortment H5N6 viruses that
contain the internal genes fragments from H9N2 [Figure 6 A], two
strains isolated during 2017 which are of the G1 genotype, and six
strains isolated during 2018. These last six strains comprise three
strains with the G1 genotype and three with the G1.4 genotype, and
differed from G1 due to a PA gene from other AIVs. We propose that the
virus can move from poultry to humans after new reassortment. The LPAIV
provides the gene segments for the reassortment event, which is a
problem that cannot be ignored. The current epidemic strain is of the G1
genotype, which is of great importance because G1 can also infect
humans. From our mice experimental data, all of the strains are H5N1,
except one, three H5N6 belong to the G1, G1.2, and G1.4 genotypes, and
show differences virulence in mice. Apparently, one strain is unable to
represent all of the genotype fully, but what caught our attention is
that H5N6 shows high virulence, which potentially increases the risk of
infection to humans.
Waterfowl have long been considered the natural reservoir of avian
influenza, and our research results show that the duck, goose, and wild
bird waterfowl contain numerous genotypes, respectively 8, 3, and 3
[Figure 6B]. Two major epidemic
genotypes, G1 and G2.1, which are distributed in the three kinds of
waterfowl. There were no G2.1 genotypes in human and chicken sources,
and we only found H9N2 gene
reassortment strains isolated from
chicken, people, and the environment. We speculate that the virus could
have spread from ducks to chickens with H9N2 reassorting and the
production of the G2.2, G3.1, and G3.3 new genotypes. The virus could
then have spread to people through chickens or the environment. G1.4 and
G3.2 are distributed only in ducks, humans, and the environment, and
from ducks or the environment can spread to people [Figure 6B]. The
pandemic genotypes G1 and G2 are distributed in wild birds, which
probably means that the virus continues to circulate in wild birds and
waterfowl. The contact between waterfowl and landfowl mainly occurs in
LPMs, which are the main place for virus reassortment and cross-host
transmission of avian influenza. Both previous research and the work of
our group confirm this scenario.
H5N1 and H5N6 can be highly pathogenic and induce high mortality in
chicken. Vaccination is one of our main measures against the highly
pathogenic avian influenza(Alarcon et al., 2018; Arai et al., 2019; Shi
et al., 2018). The Chinese vaccine is upgraded continuously, following
the evolution of the influenza virus, and the vaccine can protect
poultry effectively, reducing the virus isolation rate greatly. However,
there are still outbreaks of HPAIV every year, according to statement
from the Ministry of Agriculture. Our laboratory has received from the
farm of clinically suspected cases
of AIV. Through analysis, we confirmed that the part of the clinically
suspected case really is HPAIV, and most isolated strains from the
clinically suspected cases are reassortment viruses, which may explained
why there are still bits of outbreaks of HPAIV. Some may unuseed
vaccines,and these reassortment strains may escape the vaccine’s immune
response. A large number of reassortment viruses can be detected each
year. And the vaccine obviously cannot resist all of the new
reassortment viruses. Even so, occasional outbreaks can be addressed by
rapid culling, disinfection, and landfills, and the effect of preventing
the spread of the virus is also significant. Thus, we need to raise
enough awareness about the occasional outbreak, and only quick and
effective prevention measures can prevent a possible outbreak.