6. Conclusions and Future Perspectives
Despite much effort in studies and preparedness from deadly viruses including HPAI, SARS and MERS outbreaks, the unexpected 2009 H1N1 strain and 2019 SARS-CoV-2 quickly spread worldwide and became pandemics in 2009 and 2020, respectively. Although H5 and H7 vaccines, either monovalent or bivalent forms, are used for protection of domestic birds [252], the ever-changing and highly reassorting HPAI H5 has continued to cause outbreaks in domestic birds and to be detected in dead wild birds [24]. HPAI H7N3 in poultry has been reported until now [24]. HPAI H5N6 human cases [253] and HPAI H7N9 human cases [254] were still reported in 2019. Several avian IAVs including H7N9, HPAI H5NX (X, NA subtypes) and H9N2 sporadically cross species to infect humans. MERS-CoV is still causing an endemic in dromedary camels and human infections [8] and SARS-like CoV is still circulating in animals, notably in palm civets and raccoon dogs [100]. Thus, it is very important to understand how a new pandemic arises.
Historical data indicated that CoVs, which infect humans by using a specific host protein as a primary receptor, including HCoV-229E (APN), HCoV-NL63 (ACE2), SARS-CoV (ACE2), SARS-CoV-2 (ACE2) and MERS-CoV (DDP4), originated from bats, whereas HCoV-OC43 and HCoV-HKU1, which use Neu5,9Ac2, originated from rodents [255]. In contrast, IAVs infecting humans using sialyl glycans are thought to have originated from wild birds. The finding that some H16 HAs isolated from wild birds contain Y98F, being different from HAs of other Sia-binding IAVs but the same as bat non-Sia-binding HAs of H17 and H18 IAVs, and the finding that there are bat α2,3Sia-binding IAV isolates that are phylogenetically different from other identified IAVs with HA closest to mallard H9 viruses indicate the possibility of cross wild bird-to-bat and/or bat-to-wild bird transmission of IAVs [64]. Studies on evolutional changes of wild bird IAVs and bat IAVs may lead to clarification of the host origins of IAVs. An understanding of viral adaptation in the wild animals to recognize distinct receptors may lead to the establishment of strategies for efficient control of CoVs and IAVs.
Data from genome analysis of the past three IAV pandemics, except for the first identified 1918 H1N1 pandemic that remains a mystery, demonstrated that a pandemic emerged from viral reassortment with a human virus gene segment(s) and a nonhuman HA gene segment producing a major viral antigen [19]. Extensive studies on receptor binding specificities of IAVs from different hosts [19,256] have indicated that a nonhuman HA gene must acquire mutations to recognize α2,6Neu5Ac for efficient human-to-human transmission, leading to the development of several techniques for monitoring and assessing IAV pandemic potential, including a viral NA-based assay [257], a glycan microarray assay [258], an evanescent-field-activated fluorescence scanner type glycan array [259] and a glycan strip test [260]. However, occasional direct pig-to-human [261] and human-to-pig [262] transmission of swine and human IAVs, respectively, both of which preferentially bind to α2,6 sialyl glycans, and some avian IAVs, including HPAI H5N1 [263], H7N9 [264], H9N2 [265], H7N2 and H7N3 [266], that were reported to have increased binding to human-type α2,6Neu5Ac have not yet caused a pandemic. In addition to monitoring the α2,3 to α2,6 binding shift of HA, a simple test for monitoring shifts in other factors, such as PB2, should be developed for monitoring the situation of a virus with pandemic potential.
Different from IAVs having a segmented (-)ssRNA genome, CoVs have a nonsegmented (+)ssRNA genome. SARS-CoV and MERS-CoV originated through recombination in bats [267] and cross-species transmission to intermediate hosts, palm civets and dromedary camels, respectively, resulting in transmission to humans. Both palm civet SARSr-CoV and human SARS-CoV and dromedary camel MERS-CoV and human MERS-CoV have 99.8% nt sequence identity [217,268]. In contrast, a nonhuman virus that is almost identical to SARS-CoV-2 has not been identified so far, suggesting no widespread infections of a nearly identical SARS-CoV-2 in natural or intermediate hosts [269]. Based on current data, SARS-CoV-2 might have emerged from a quadruple recombination by which Yunnan bat RaTG13 (96.1% nt identity with SARS-CoV-2) might be its genome backbone [226], Yunnan bat RmYN02 (93.3% nt identity) probably gave the long replicase gene (1ab gene, sharing 97.2% nt identity) [133], pangolin SARS-like-CoV-2/Guangdong might have provided an RBD motif-coding gene (97.4% aa similarity) [231], and an unidentified bat virus might have donated a gene region coding a multibasic (furin, PRRAR motif) cleavage site [133]. Similarly, the 2009 H1N1 pandemic is a quadruple reassortant IAV that acquired gene segments from human IAV (PB1 gene), avian IAV (PB2 and PA genes), classical swine IAV (H1, NP and NS genes) and Eurasian avian-like swine IAV (N1 and M genes) [4].
The ability of a virus to adapt to a human environment, including human immunity and drugs, enables it to seasonally circulate in humans. Highly transmissible SARS-CoV-2, which has the same ACE2 receptor as that of HCoV-NL63, may continue to be a seasonal CoV and may undergo recombination with HCoV-NL63 during mixed infection in the same cell. While there is no vaccine for and no specific drugs for treatment of mild common cold disease caused by HCoVs, many efforts are being made to develop both vaccines and drugs for the pandemic SARS-CoV-2. Influenza caused by seasonal IAVs can be prevented by yearly vaccination and can be treated by FDA-approved anti-flu drugs, which are divided into four groups: M2 inhibitors, a PA inhibitor, a PB1 inhibitor and NA inhibitors [270]. Nonetheless, highly mutable IAVs that possess RNA genome segments and RNA polymerase without proofreading, different from the CoV RNA genome and RNA polymerase with proofreading, have continued to circulate in humans for more than one hundred years. Structural analysis, studies on receptor binding specificity and studies on inhibition of infection by synthetic sialylglycopolymers [19,271] have suggested that there is an invariant receptor binding site (RBS) on viral HA spikes that is critical for virus binding to a human-type receptor for infection. This easily reachable drug target that is abundant on the viral envelope is a promising target for the development of a universal and permanent anti-influenza drug against human-adapted viruses of H1–H16 HA subtypes. Recently, 6SLN-lipo PGA [270,271] was shown to be effective when administered alone and to have a synergistic effect when combined with an NAI drug against both pandemic and seasonal influenza viruses. In the case of CoVs, the RBS could be a potential drug target. However, CoVs recognize different receptors including protein receptors, CEACAM1a, NCAM, DDP4, APN and ACE2, and saccharide receptors, sialylated or non-sialylated saccharides. Some CoVs also recognize a co-receptor/attachment factor by a different binding pocket of the S protein and some βCoVs also carry an HE spike protein binding to an O-acetylated Sia receptor (Table 2). Thus, for the design of an anti-CoV against the RBS, consideration must be given to the role and importance of each receptor and to the use of two inhibitors in combination or the possibility of generating a single compound with two different inhibitory sites.
Finally, studies to understand how human viruses emerge, to understand the pathogenesis of diseases, and to produce effective, safe and permanent (if possible) vaccines/drugs are essential for virus control and eradication. However, the most important thing is creating awareness about the results of human intrusion and disturbance of wildlife in order to minimize or eliminate direct contact of domestic animals/humans with wild animals for prevention of the next emerging disease.