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May 25 15 tweets 6 min read Read on X
In new study led by @bdadonaite, we measure how all mutations to H5 influenza HA affect four molecular phenotypes relevant to pandemic risk:


Results can inform surveillance of ongoing evolution of H5N1. biorxiv.org/content/10.110…
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To measure how all HA mutations affect those phenotypes, we created pseudovirus libraries of HA from WHO clade 2.3.4.4b vaccine strain.

Pseudoviruses encode no genes other than HA, so can only do a single cycle of infection making them safe for biosafety-level-2. Image
First, we measured how all mutations affected HA-mediated cell entry, which is essential for viral fitness

See heatmap below, which is easily visualized interactively at

Some sites constrained (orange); others w many well tolerated mutations (white/blue) dms-vep.org/Flu_H5_America…
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Here is constraint (average cell entry effect of mutations at each site) on HA structure.

These measurements define ideal targets for new approaches (eg ) to develop antibodies targeting constrained regions of virus. biorxiv.org/content/10.110…
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Next, we measured how mutations affect HA’s usage of a2-6 vs a2-3 sialic acid receptors (thanks @PeacockFlu for idea how to do this)

Important because human-transmissible viruses use a2-6.

Below are mutations that improve a2-6 usage, and should be monitored for in surveillance Image
Airborne-transmissible influenza viruses tend to have higher HA stability, so we also measured how all mutations affect stability.

As seen below, stability enhancing mutations tend to be located in helices in fusion machinery and interfaces between head & stalk domains. Image
In addition to relevance of stability-enhancing mutations for pandemic-risk surveillance, these mutations could be useful to introduce into vaccine immunogens as stabilizing mutations have improved immunogenicity of other viral vaccines.
We next measured how all HA mutations affected neutralization by sera from vaccinated mice (from @ScottEHensley) or ferrets (from Richard Webby).

This is important because WHO recommends candidate vaccine strains to H5, which may need to be updated as virus evolves.
Below are sites in HA where mutations reduce neutralization by mouse or ferret sera (see interactive plots w per-mutation effects)

These data make it possible to rapidly identify new viral mutants that are mismatched to current candidate vaccine strains. dms-vep.org/Flu_H5_America…
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To use these experimental datasets to inform surveillance, Jordan Ort & @LouiseHMoncla to integrated them into an interactive Nextstrain tree () that allows you to color nodes by phenotypes (see dropdown at left)nextstrain.org/groups/moncla-…
Such trees make it possible to immediately identify when new viral mutants have potentially relevant HA phenotypic changes; for instance, see below for examples of mutations in dairy cattle cluster that reduce neutralization, or viruses w increased HA stability. Image
The datasets are so rich that we recommend viewing interactive plots rather than static figures:

- Homepage w all data:

- Data mapped on HA structure:

- Data mapped on phylogenetic trees: dms-vep.org/Flu_H5_America…
dms-viz.github.io/v0/?data=https…
nextstrain.org/groups/moncla-…
In addition, we have made a page to help convert between the many confusing numbering schemes used to name HA mutations:

(Throughout we use H3 numbering, but other studies use other schemes)dms-vep.org/Flu_H5_America…
Finally, as we discuss in paper, our ability to safely do deep mutational scanning of H5 HA enabled by switch from live-virus to pseudovirus. I thank @mlipsitch & others for thoughtful discussion that prompted switch; we welcome continued input on those aspects of our work. Image
Thanks to all who contributed to this study: @bdadonaite, @Jenny_Ahn0, Jordan Ort, Jin Yu, Colleen Furey, @anniedosey, Will Hannon, Amy Baker, Richard Webby, @KingLabIPD, Yan Liu, @ScottEHensley, @PeacockFlu, @LouiseHMoncla

Pre-print is here: biorxiv.org/content/10.110…

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More from @jbloom_lab

Apr 20
In new study led by @bblarsen1 in collab w @veeslerlab @VUMC_Vaccines we map functional & antigenic landscape of Nipah virus receptor binding protein (RBP)


Results elucidate constraints on RBP function & provide insight re protein’s evolutionary potentialbiorxiv.org/content/10.110…
Nipah is bat virus that sporadically infects humans w high (~70%) fatality rate. Has been limited human transmission

Like other paramyxoviruses, Nipah uses two proteins to enter cells: RBP binds receptor & then triggers fusion (F) protein by process that is not fully understood
RBP forms tetramer in which 4 constituent monomers (which are all identical in sequence) adopt 3 distinct conformations

RBP binds to two receptors, EFNB2 & EFNB3

RBP’s affinity for EFNB2 is very high (~0.1 nM, over an order of magnitude higher than SARSCoV2’s affinity for ACE2) Image
Read 12 tweets
Mar 5
Over 4 yrs after being first to publicly release SARS-CoV-2 genome, Yong-Zhen Zhang just published large set of viral seqs from first stage of COVID-19 outbreak in China


He uses data to suggest scenarios re early outbreak & root of viral phylogenetic tree academic.oup.com/ve/advance-art…
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Zhang recruited nearly all COVID-19 patients hospitalized at Shanghai Public Health Center in first 2/3 (Jan-Sep) of 2020.

The largest source of Shanghai patients in Jan/Feb 2020 was imported cases from Wuhan or elsewhere in Hubei, thereby providing window into Wuhan outbreak. Image
Overall, Zhang obtained 343 near-full-length SARS-CoV-2 sequences from 226 distinct patients, including 133 sequences from samples collected no later than Feb-15-2020.

A phylogenetic tree showing these sequences is below. Image
Read 11 tweets
Feb 7
In new study led by Caleb Carr & @khdcrawford, we measure how all mutation to Lassa virus glycoprotein complex (GPC) affect cell entry & antibody escape

Results show how prospective assessment of effects of mutations can inform design of countermeasures
biorxiv.org/content/10.110…
As background, Lassa virus causes of thousands of deaths each year, mostly from spillovers from its rodent host, but there is occasional human-to-human transmission.

Lassa is biosafety-level-4 priority pathogen, & efforts are underway to develop vaccines & antibody therapeutics.
We used pseudovirus deep mutational scanning to study effects of nearly all 9,820 amino-acid mutations to Lassa’s GPC at biosafety-level-2 by making genotype-phenotype linked libraries of lentiviral pseudotypes
blog.addgene.org/viral-vectors-…
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Read 18 tweets
Jan 17
Here is my brief analysis of Dec-28-2019 SARSCoV2 submission to Genbank.

This analysis supports my conclusion to WSJ () that this submission does not tell origin of virus, but does show sequence known to Chinese Academy of Sciences weeks before released wsj.com/politics/natio…
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Here is link to my full analysis:

See also images of the same posted below (although it's probably just easier to click on link above and read HTML). github.com/jbloom/SARS2_2…



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I also don't think Genbank/NCBI could have reasonably known at time that this sequence was so valuable given that Chinese govt did not announce they had sequence or had submitted it, and Genbank receives vast numbers of submissions.
Read 4 tweets
Dec 17, 2023
In new study led by Frances Welsh, we map how mutations to influenza affect neutralization by antibodies from humans of various ages

We find differences in mutation effects among age groups

Virus has evolved especially to escape antibodies of teenagers

biorxiv.org/content/10.110…
As background, human influenza constantly evolving. So people exposed to different strains, depending on their age & idiosyncratic history of infection/vaccination.

Different exposure histories cause people to make antibodies w different specificities

rupress.org/jem/article/21…
How does this person-to-person heterogeneity in antibody specificity affect influenza evolution?

That’s question we set out to answer

We used deep mutational scanning to measure how H3N2 HA mutations affect neutralization by serum antibodies from children, teenagers, and adults
Read 11 tweets
Nov 29, 2023
I wanted to highlight this pre-print by David Ho’s group on the neutralizing antibody response to new (XBB.1.5-based) COVID vaccine booster, as it illustrates some points related to paradigm of updating SARS-CoV-2 vaccines to keep pace w viral evolution.
biorxiv.org/content/10.110…
Recall original COVID vaccines worked very well against early SARS-CoV-2 strains

Unfortunately, virus has been evolving, so antibodies elicited by that vaccine don’t neutralize newer viral variants very well

(Other human CoVs also evolve same way: ) journals.plos.org/plospathogens/…
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So in fall 2022, new booster was made that mixed new (at time) BA.5 variant & original strain. Hope was to boost neutralization of new variants.

Unfortunately, only sort of worked. Titers did go up, but not a relatively greater increase for new variants. Image
Read 15 tweets

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