Discussion - 2019-nCoV genetics

[JJackson]

For those of us with an interest in viral genetics I have started this thread to look at the little data that is now available. What follows is not my analysis but based on the work posted on the Virological.org forum.

1] The consensus is that much of the sequence data is unreliable due to the high number of sequencing errors.
2] Based on the more trusted sequences the MRCA (most recent common ancestor) is probably in early Dec. 2019 giving a recent single common ancestor for all sequences. If accurate this means it probably has not been circulating below the radar for a while.
3] Due to the recent MRCA the cladogram is boringly flat with only a one or two AA variance from WH01 and most being identical. Where there are changes they seem random with no evidence of host adaption.

A note on the Virological site. If you followed the H7N9 discussion thread you may remember Andrew Rambaut from Edinburgh Uni. who posted really useful phylogenetic trees and analysis on the University's Epidemic site (which had sections on MERS, Ebola and flu). When this outbreak occurred I checked the site only to find it had disappeared but Andrew had reappeared as Administrator of Virological which at that time only had a couple of threads. In the main discussion thread I posted a speculation this was a Beta CoV before it was announced, I would not normally have done this based on a single media report but the article was by Lisa at Cidrap and she was quoting Marion Koopmans of Erasmus. I trust MK not to have made the comment without good reason and LS not to use it without reason to think MK had good reason for saying it. Which brings me back to Virological where MK & AR are contributing and which was used to post the first open source sequence. Currently some sequences are being deposited at GISAID and others at Genebank.

 

[gsgs]

there was speculation about a lab-release , biggameindia zerohedge , are the mutations and comparisons with older bat-viruses matching or are there timely gaps , has this been checked

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I'm currently downloading and aligning coronaviruses from genbank, got 388 , so slow with mafft

 

[JJackson]

I am learning as I go as I have not looked at CoVs before. What I have gathered is that a fair bit of research was performed in to SL SARS which are the group of beta CoVs found in the host reservoir that forms the genetic pool from which SARS emerged. Much of the RNA is conserved with 95%+ homology but like flu there are areas where that drops to 85%ish. One area that seems to be a focus of interest S 153syn which sits in the S2 binding pocket making it a potential drug target which, if I understood the TWiV you linked to, is an ACE2 receptor. This pocket is part of the low homology zone and includes the primary antigenic site making life harder for the immune system. Much of this may be my poor understanding of what I read as I am getting out of my comfort zone with these papers.

 

[gsgs]

we need this RaTG13 virus, which is by far the closest.
Allegedly from Rhinolofus affinis from Yunnan


https://www.biorxiv.org/content/10.1...01.22.914952v2


Data Availability statement. Sequence data that support the findings of this study
have been deposited in GISAID with the accession no. EPI_ISL_402124 and
EPI_ISL_402127-402130.

https://en.wikipedia.org/wiki/Interm..._horseshoe_bat

---------------------------------------------------------------------------------
Yen Shu Chen • 3 days ago
The authors did not share (no GenBank/GISAID accession number are
provided) the genome sequence of the critical bat-CoV that represents a
close relative to human 2019-nCoV.
No way to access/reproduce/further use their result. Do scientific journals accept such practice?
-------------------------------------------------------------------------------
reuns Yen Shu Chen • 2 days ago • edited
Exactly, there has been thousands of bats coronavirus sequenced since the Sars epidemic,
it is not unusual that most of them are just lost into some dusty lab, but it is weird they
didn't upload thisone since it is the most astonishing result of their study, and the exact
genome might contain some useful information on how bat viruses can mutate and
contaminate human. Also note that the paper is from Wuhan's institute of virology, probably
the same lab which discovered the aforementioned bat coronavirus
-----------------------------------------------------------------------------

 

[gsgs]

I finally got it aligned etc. , 395 coronavirus genomes from genbank, mostly SARS.
file corona20.c6 , 16MB , someone wants it ?
The recombinations are not so clear and there could be variations of the mutation rate in the 30000 nucleotide genome
I made these charts, #mutations per 100 nucleotides: http://magictour.free.fr/WUH01.GIF
http://magictour.free.fr/WUH11.GIF

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the fasta was 1.2GB, now downloading the whole genbank records with the dates etc.

 

[JJackson]

gs Recombination seems to be a feature of this virus, along with deletions and insertions. There has been an interesting conversation developing over at Virological relating to the dangers of using whole genome homology or even single gene homology to achieve the true proximity of isolates. Large recombination events causing poor homology overall while very high homology remains across the unaffected sections of RNA.

Do you have a sample collection date for RaTG13? The very high homology across the Spike gene is at odds with everything else except nCoV2019 including all other bat SL CoVs given that this is probably the least conserved region, unless the sample was very recent, I do not see how it has maintained its sequence so faithfully nor why this outlier has given its S gene genetics to nCoV. Is there something specific to this unique Spike sequence that makes it well adapted to infect humans? The SL CoVs generally are well know for there ability to spread to other mammalian host (civet, racoon dog etc.) so it seems unlikely that these two Chinese culinary favourites have not presented SL CoVs to humans since SARS yet this atypical sequence has very successfully made the jump and seems to have little difficult binding to our ACE2 receptors (assuming that is what they are using for access this time around).

Edit:
I found the RaTG13 date and it was 2013 so several years of drift.

 

[gsgs]

2013 is still 7 years ago, surprisingly long for the 96.5% similarity. Compared with the other viruses.
Suggesting lots of bat-coronavrus diversity ...
I don't know about the different proteins but read about the recombinations and different mutation rates.
That's why I made the pics with mutationrate over the 30000-genome.

The region ~12000-~20000 looks suitable for mutation-timing-comparison

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where did you find 2013 ? It's not in the paper. Well, the 13 in RaTG13 may stand for 2013


> RaTG13 which we previously detected in Rhinolophus affinis from Yunnan Province showed

I couldn't find a current authors in a related reference

Yang, L. et al. Novel SARS-like Betacoronaviruses in Bats, China, 2011.
Emerg Infect Dis 19, 989-991, (2013)

Hu, B. et al. Discovery of a rich gene pool of bat SARS-related
coronaviruses provides new insights into the origin of SARS coronavirus.
PLoS pathogens 13, e1006698, (2017)

Wang, N. et al. Serological Evidence of Bat SARS-Related Coronavirus
Infection in Humans, China. Virol Sin 33, 104-107, (2018)

 

[Emily]

There is an interesting post at Virological.org, “not snakes v2”, that discusses fungi and SARS, MERS, and nCov-2019. They joke that in spite of the high CAI values that there is no significance to this.

I'm wondering now.

Widespread Bat White-Nose Syndrome Fungus, Northeastern China

Do Viruses Exchange Genes across Superkingdoms of Life?

 

[gsgs]

here is another paper :
https://www.biorxiv.org/content/10.1...249v1.full.pdf
> The BatCoV RaTG13 sequence was downloaded from the GISAID BetaCov 2019-2020 repository

IMO it's a shame, that this important sequence is not public and at genbank.
When it's at GISAID, it may only be revealed to other GISAID member,
that's not really "publicly available", as they claim.

------------edit-------------------
no recombination BaTG13 -- 2019-nCoV

 

[JJackson]

This is interesting https://www.bilibili.com/read/cv4457676/ it is in Chinese but this worked for me https://translate.google.com/transla...2Fcv4457676%2F

This is the first I have seen on lab tests with live nCoV virus.
They look at two questions
1] Is the ACE2 receptor, used by SARS, also used here. They used HeLa cells modified to display various ACE2 receptors from humans, bats, civets, pigs and mice - the virus grew in all but mice so mice are probably out as a lab animal unless a there is a linage with modified ACE2s.
2] They also looked at serum treatments and both human and horse anti-sera worked well and could be a useful therapeutic at least as a stop gap measure.
This graphic nicely illustrates just how far RaTG13 has wondered from the pack (it is rooted against nCoV2019) and helpfully provides a nucleotide and AA scale across the top. ZC45 seems to share a high homology across ORF1a but reverts to the pack for the Spike protein. I assume that ORF just stands for Open Reading frame but it seems odd that it is not given a designation based on a protein produced as they account for 2/3 of the genome.

I can not find the paper I found RaTG13 was from 2013 but it definitely gave that as the date and stated the host as horse shoe bat. For RaTG13 to hold such a high % homology for so long makes me wonder if there is another host species in which this RBD configuration needs to be conserved and nCoV came from this source and RaTG13 is a reintroduction to bats and ZC45 is a recombination event. As is often the case in wild animal sampling there are just too few relevant sequences to be sure we have any real understanding of the viral genetic dynamics much of what we do have is due to post SARS investigation and fairly dated given the speed of viral evolution.

 

[JJackson]

This is a useful visualisation tool https://nextstrain.org/ncov

 

[gsgs]

how are these pictures called ? Also 3dim ? I didn't first understand it
and did the same in multi-pics, one per chart..
It looks as if the mutation rate is just different in different areas
I figured out that RatG13 is at GISAID
can we email or pm seems that i can't pm or send visitor message
You might apply some function to account for the mutationrates per region
Isn't it likely that China has many more bat-sequences with
multiple strains but only academics publish because it's good for their career
I complete my genbank downloads ... still reorginising data with my old flu-tools

 

[gsgs]

https://flutrackers.com/forum/forum/...ations-cloning

[after WIV1 in 2012]
The sampling of this bat cave in Yunnan continued for another 5 years. From these samples,
the research team isolated 3 live viruses in succession, and obtained the full-length genomic
sequences of a total of 15 bat SARS-like coronaviruses. Surprisingly, the 15 strains contained
all the genome components of the SARS virus.
Hu Ben, an assistant researcher at the Wuhan Institute of Virology, Chinese Academy of Sciences:
Shi Zhengli
...
fuchsia line
the highest similarity between SARS-like coronaviruses found in bat caves and their respective
genes is above 97%,
After 13 years of virus tracing, the origin of the SARS virus was finally found.

[ one of those Yunnan cave viruses must have been RaTG13 , 96% similar to 2019-nCoV ]

 

[gsgs]

http://virological.org/t/ncov-2019-s...ic-dataset/362

So far, the nCoV-2019 has been reported to share 96% sequence identity to the RaTG13
genome (EPI_ISL_402131). However, the S1 Receptor Binding Domain (RBD) of the nCoV-2019
genome was noticeably divergent between the two at amino acid residues 350 to 550 – Figure 1A.
We aimed to identity coronaviruses related to nCoV-2019 in viral metagenomics datasets available
in the public domain. In a recently published dataset describing viral diversity in
Malayan pangolins (PRJNA573298 10) we used VirMAP 11 to reconstruct a coronavirus genome
(approximately 84% complete from samples SRR10168377 2 and SRR10168378 1) that
shared 97% amino acid identity across the same RBD segment – Figure 1B. This result
indicates a potential recombination event for nCoV-2019.

[these are amino-acids, but what about nucleotides ?
I remember the amino-acid conservation in inner segments of flu-A in mallards]

pangolin=Schuppentier , https://en.wikipedia.org/wiki/Pangolin

 

[bertrand789]

La m?me source a produit aussi ceci : https://www.nature.com/articles/s41586-018-0010-9

 

[Vibrant62]

I have a question - how much reassortment / recombination goes on between CoV viruses of different clades. For example if 2019 nCoV meets MERS, is there a risk of reassortment?

 

[bertrand789]

des lieux ou des porcs et des humains cohabitent en Chine cela me semble exister d'une part , d'autre part on ne sait rien de ce qui se passe sur les porcs sauvages chinois , donc les sangliers ...

 

[Pathfinder]

Mining coronavirus genomes for clues to the outbreak’s origins

By Jon CohenJan. 31, 2020 , 6:20 PM
...
“One of the biggest takeaway messages [from the viral sequences] is that there was a single introduction into humans and then human-to-human spread,” says Trevor Bedford, a bioinformatics specialist at the University of Washington and Fred Hutchinson Cancer Research Center.
...
The longer a virus circulates in a human populations, the more time it has to develop mutations that differentiate strains in infected people, and given that the 2019-nCoV sequences analyzed to date differ from each other by seven nucleotides at most, this suggests it jumped into humans very recently. But it remains a mystery which animal spread the virus to humans.
...
According to Xinhua, the state-run news agency, “environmental sampling” of the Wuhan seafood market has found evidence of 2019-nCoV. Of the 585 samples tested, 33 were positive for 2019-nCoV and all were in the huge market’s western portion, which is where wildlife were sold. “The positive tests from the wet market are hugely important,” says Edward Holmes, an evolutionary biologist at the University of Sydney ...
...
Yet there have been no preprints or official scientific reports on the sampling, so it’s not clear which, if any, animals tested positive. “Until you consistently isolate the virus out of a single species, it’s really, really difficult to try and determine what the natural host is,” says Kristian Andersen, an evolutionary biologist at Scripps Research.
...
It’s not just a “curious interest” to figure out what sparked the current outbreak, Daszak says. “If we don't find the origin, it could still be a raging infection at a farm somewhere, and once this outbreak dies, there could be a continued spillover that’s really hard to stop. But the jury is still out on what the real origins of this are.”

https://www.sciencemag.org/news/2020/01/mining-coronavirus-genomes-clues-outbreak-s-origins

 

[alert]

I have a question - how much reassortment / recombination goes on between CoV viruses of different clades. For example if 2019 nCoV meets MERS, is there a risk of reassortment?

Yeah, I'd think that's one of the real dangers. Because MERS is much more severe, and you might get a resultant virus that spreads as easily as 2019 nCoV but has the severity of MERS.

I'm also concerned about what happens if someone gets co-infected with 2019 nCoV and one of the four seasonal human coronaviruses, which seems more likely than a MERS co-infection.

 

[bertrand789]

L'un des probl?mes me semble que lors de la fermeture des proc?dures judiciaires ont du ?tre ouvertes. Je crains que certains ?levages ou fournisseurs d'animaux interdits ont alimente x lieux. En revanche, je suis s?r qu'ils cherchent et vu leurs comp?tences, ils vont trouver. Pour faire entrevoir de fa?on imag?e, le dossier march? global, en Chine, c'est comme celui de la vente d'arme au U. S. A. En tout cas entre le dossier peste porcine africaine et celui ci cela doit ?tre lourd... Ils doivent ?tre f?licit?s et non critiques, de mon point de vue... Mais apr?s la phase de crise, il faudra vraiment trai