The Covid Genome

Question 1

Genomics of Covid-19. Exeter contributing to national effort to sequence genomes of as many isolates of the virus as possible. What will this help with?

Answer 1

Developing vaccines, inform development of testing diagnostics kits and insight

Question 2

Coronavirus officially known as

Answer 2

SARS-CoV-2

Question 3

What does SARS-CoV-2 cause?

Answer 3

COVID-19 disease

Question 4

Genome composition of SARS-COV-2

Answer 4

single stranded RNA virus genome consisting of ssRNA in the positive orientation

Question 5

Genome consist of

Answer 5

ss RNA (under 30kb in length)

Question 6

Genome consists of several

Answer 6

open reading frames translated into polypeptide protein products

Question 7

What became available at the end of 2019?

Answer 7

First genome sequences of isolates of the virus from China

Question 8

What were the researchers next step?

Answer 8

Researchers looked at the whole genome sequences and compared to whole genome sequences of other viruses, researchers were able to come up with a phylogenetic tree

Question 9

Coronavirus probably circulating in

Answer 9

bats as main natural reservoir but it probably didn’t come directly from bats

Question 10

Background to the development of coronavirus: why is it likely that coronavirus didn’t come directly from bats?

Answer 10

In December, bats hibernate. No bats in Huanan seafood market (but many other mammal species which could have become an intermediate host). Under 90% identity to closest bat virus

Question 11

SARS and MERS

Answer 11

emerged via intermediate hosts

Question 12

SARS intermediate host

Answer 12

association with palm civets (virus transferred form bats to palm civets and then to humans)

Question 13

MERS intermediate host

Answer 13

via camels to humans

Question 14

Current method of COVID-19 testing

Answer 14

swab and perform PCR

Question 15

from comparing the first few genome sequences of the current pandemic (COVID) against other genome sequences of previous outbreaks (SARS + MERS) and other viruses from bats you can see that some parts of the viral genome are

Answer 15

highly conserved along most maybe all coronaviruses (whereas some are more variable)

Question 16

It’s essential to design

Answer 16

PCR primers that target the more conserved (less variable) regions of the genome

Question 17

An example of highly conserved region which could be targeted with a detection assay which relies on PCR

Answer 17

1b (in this diagram) seems quite highly conserved amongst different viruses within this group (in addition to places on the other end of the genome which show relatively high levels of consensus)

Question 18

Example of study which did this

Answer 18

Detection of 2019 novel coronavirus (2019-nCoV) by real time RT-PCR = identified regions of the genome which seemed highly conserved and designed PCR primers against them. some of these primers are now recommended by Public Health England

Question 19

By 05/04/202

Answer 19

up to 3,000 genome sequences available for the virus.

Question 20

UCSC genome browser

Answer 20

has a genome browser for the SARS-Cov-2 viral genome

Question 21

UCSC genome browser. The SARS-CoV-2 reference genome in this case was

Answer 21

originally isolated virus from one of the first patients from China

Question 22

UCSC genome browser. what can you see

Answer 22

You can see the positions of the genes (protein coded genes) which are annotated. Target sites of the PCR primers can be seen, used to test and detect the virus

Question 23

UCSC genome browser. where are PCR primers targeting?

Answer 23

Many PCR primers are targeting the 1b highly conserved region of the genome Others target other parts of the genome (also highly conserved)

Question 24

UCSC genome browser. Towards the bottom of the screen you can see the variation between different isolates of the virus

Answer 24

Each of these rows is the genome sequence of 1 isolate of the virus. Where there’s a difference in the genome of that isolate compared to the reference there is a dot. Many dots scattered around; large number of isolates which have a variant in the same region of the genome (long lines down). As the virus passes through hosts and replicates its accumulating mutations

Question 25

What do you not want to see in the highly conserved regions of the genome

Answer 25

YOU do not want to see many differences/ variants (dots) in the highly conserved regions of the genome (targeted by our PCR primers – they could fail to target the viral genome if too many variants)

Question 26

Genomic epidemiology of novel coronavirus – Next strain Resource

Answer 26

good for exploring genomic data from current pandemic. Data gathered by the GISAID consortium. Over 3,000 genome sequences of CoV-SARS-2 in the data set

Question 27

Historically what did GISAID focus on

Answer 27

strains of influenza but now collecting genomic data on current pandemic

Question 28

Next strain resource has the

Answer 28

latest data and analysis

Question 29

Next strain resource shows diversity

Answer 29

o Genetic diversity (sequence diversity) o Shows the 30kb of the genome o Height of spikes shows how much genetic diversity exists at each position  most have next to none (most of the genome is completely conserved) but some hotspots along the genome – considerable amount of sequence variation because of mutations during replication of the virus during the pandemic

Question 30

Next strain resource shows phylogenetic tree

Answer 30

o Phylogenetic tree of the 3,000 viral genome sequences which are Colour coded according to geographical origin

Question 31

Next strain resource shows - Geographical map of transmission

Answer 31

o Press play – reconstruction of outbreak according to timeline of transmission

Question 32

Next strain resource shows filter by country

Answer 32

533 genome sequences from the uk o Able to tell you when it was first sequenced and where

Question 33

Next strain resource can explore patterns further

Answer 33

o Go back to main page o Look at situation reports o Can see analysis = Recently another 16 genome sequences have been deposited from Japan, 10 of the 16 come from patients who were on a cruise. These isolates more closely resemble isolates from Europe and North America than Japan, this reflects that the Japanese patients were infected on the cruise by other patients from Europe or North America rather than developing the infection in Japan. 2 genome sequences from Wales they don’t cluster together this suggests the virus has been introduced into wales more than once

Question 34

Exeter’s efforts

Answer 34

uni using Oxford nanopore minions to sequence isolates of the virus acquired from RD+E hospital. They got the first 4 genomes from Exeter (3rd of April 2020) Placed the Exeter isolates in the phylogeny (black circles). 2 of the isolates were genetically identical to each other so only 3 black dots

Question 35

Exeter’s efforts continued

Answer 35

Using HARVEST software to look at phylogenetic tree of several hundred UK isolates of virus. Lines indicate mutations/ variants compared to reference genome Can show single nucleotide resolution through zooming in. At particular regions there are mutations which you can see on a nucleotide level