Discussion
We investigated here the origin of SARS-COV2 virus that created pandemic
in all over the world with considerable morbidity. In near future the
chance of getting a vaccine is far from reality and much more morbidity
is expected. It is imperative that currently people must depend on the
various medicines only with a trial and error basis. To develop
effective vaccines and medicines and for testing them in animals, it is
necessary to know the origin of this virus and their intermediate host
if any existed before its emergence as a major human infecting virus.
Among
the three possibilities predicted earlier whether SARS-COV2 directly
came from RaTG13 from bat using defective entry-point residues in RBD
with poor infective power with less efficiency as it is observed in
civet cats (7) or forceful infection in mice (18) and remained silent
for long time but highly adapted to replicate with slower mutation rate
and survive in a specialized immune system of the human body.
Eventually, the entry point residues have been modified and perfected to
attain widespread infectivity; or it gained the efficient entry-point
mutations first to bind with ACE2 receptor to enter human body and then
it perfected itself to adapt with higher mutation rate and evade the
human immune system; or it entered to an intermediate human like host
from bat with defective entry-point residues and adapted long time, then
entered human host recently and survived easily with optimum mutation
fitness. In all cases, after adaptation in human host, it gained more
virulence by further substitution followed by selection pressure.
Our analysis indicates the occurrence of extremely low frequency of
SARS-COV2 mutation in the human host. Mutation frequency can be
confounded by selection and genetic drift. In optimal mutational
fitness, mutation frequency is generally biased towards nonlethal
mutations and most mutations are either beneficial or neutral, thus may
dramatically underestimates mutation frequency. In that case mutation
rate could be lowered as the deleterious mutation drives the mutation
rate lower (13). Between two models as speed vs adaptability of viral
mutation rate, here it appears that SAR-COV2 evolution fits with
adaptability model. Adaptability model states that after a long
adaptation to evade immune system, the selection pressure is relatively
low and the supply of beneficial mutation frequency is reduced, thus
population favors a low mutation rate. When the mutation reaches to an
optimum level simply because selection is acting on it long time within
the context of immune escape to reach the maximum mutation fitness (13,
14, 19, 20).
If SARS-COV2 has come directly from bat as it is presumed, it would take
a very long time to evolute as a present-day SARS-COV2 virus in the
human host. Only assumption that permits this kind of viral association
in the human respiratory tract by staying as a silent virus and then
gained the virulence after a long time of adaptation. In the last 6
months the emergence of a new strain with more infective power has been
demonstrated (10). Such a creation of a strain with more infective power
also suggests that SARS-COV2 might not reside in the human host very
long time without revealing its existence even in very mild form when
human immune system tend to defeat its very existence.
However, our analysis has some limitations. It is unknown why the
mutation rates are almost double (4.89nt) in April than other previous
months. A biased sampling of a particular variant strain could represent
repetitively over other low mutating strain or inclusion of a single
genome consists of a 17base deletion or as expected by increasing
generations in April than previous months for widespread infectivity.
Although, the continuation of this increasing trend could not be
verified due to unavailability of SARS-COV2 genomic sequences beyond
April. Also, we wanted to assess here the average mutation rate in
SARS-COV2 virus and not a strain specific by assuming all strains are
capable of infecting human efficiently and undergoing substitutions to
evolute to become a better strain. We also did not separate out the
synonymous or nonsynonymous mutations although nonsynonymous mutation
selection would have been much stringent. Another important
consideration is that we did not observe any recombination or big
insertions (except one that is collected in Washington in April) in
these four months and frequent occurrence of those could increase the
mutation rate that can occur any time. However, such an event could be
very rare in an optimally mutationally fitted virus and may not add much
weightage in overall mutation rate in the long run. Lastly, we estimated
the mutation rate of SARS-COV2 in human host but extended it to
calculate the time taken by this virus from bat RaTG13. Although such an
estimation may need extensive experimental study in bat system as there
would be different selection pressure than human. Nevertheless, to take
less time to evolute in bat than human (<30years) could
presume bat system must have higher mutation rate than human which
further assume that it has to face much more challenging environment in
bat than human but that is not expected as RaTG13 is native (long time
adaptation) to bat.
Among the key entry-point residues in SARS-COV2 455L, 459Y and 500T are
same in both RaTG13 and Pan_SL_COV_GD, thus they can come from any of
them. The most important residue for SARS-COV2 interaction with human
ACE2 is K353 that binds with 501N and can evolute by conversion of D
(aspartic acid) to N (aspargine) by a single nucleotide mutation
(G>A). It is also to be noted that a single nucleotide
mutation almost gave RaTG13 a passport to infect human efficiently.
But 493Q needs mutation in two nucleotides in 1st(U>C) and 3rd codon (U>A)
sequentially either it would generate a nonsense codon (UAA). Again if
1st codon mutation occurred before
3rd codon it would code Histidine (H) by CAU. Thus,
493H carrying intermediate ancestor of SARS-COV2 virus must exists in
bat or pangolin or in any other intermediate host.
However, although a genetic drift might come into play in these
conversion from Y >H>Q but such a drift can
occur only after entering it into human or intermediate host. The silent
presence of SARS-COV2 related virus is not documented in human for long
time. Also, no evidence has supported the notion that any such primate
population are endangered/suffered due to a recent viral attack. The
mutation must be inside a host but there is a possibility that this
intermediate host no longer exists (wiped out) any more in nature or yet
to be explored. Also, with the current genomic and amino acid sequences
of SARS-COV2 having 493Q and 501N in the RBM suggests that SARS-COV2
could infect any of the primate or higher order mammals as intermediate
host having K31 and K353 residues in their ACE2 receptor gene. But till
date, none of them are shown to naturally harbor SARS-COV2 or any
closely related virus. Li et al (2004) (4) also suggested that such an
intermediate host can never be identified. Although, it is impossible to
conclude that such an intermediate host can never be found, a systematic
investigation can be continued to search for such a host.
Taken together, our analysis do not satisfy any of these conditions such
as absence of any evidence of silent presence of SARS-COV2 virus in
human for a long time that would take approximately 30 years to evolute
as a present day SARS-COV2 or very high mutation rate or a must needed
intermediate host carrying intermediate virus with 493H. Taken together,
the absence of any intermediate host or virus between bat and human and
inability to stay long time silently in human host also can lead to
believe that SARS-COV2 would have been more easier to be created
unnaturally.