#GetYourFluShot! We don’t have a SARS-CoV-2 vaccine yet, but we do have an influenza vaccine, which can play a crucial role in helping keep the coronavirus crisis from getting worse! The “flu shot” isn’t perfect (and I’ll get into why that is), but it typically is ~40-60% effective and people who get infected despite getting the vaccine often have much milder symptoms. 

This year, more than ever, it’s important to get your flu shot. COVID-19 is already sickening and killing unfathomable numbers of people and placing a huge burden on our health care system. Imagine how much worse it could be if there’s a bad flu season on top of it. I know a lot of people don’t typically get flu shots, but I hope I can convince you…

I’m going to go into a lot more detail (hopefully not more than you want…) but I will start with a kinda-TLDR about why it’s so hard to make effective flu vaccines (and why it should hopefully be easier to get an effective SARS-CoV-2 vaccine…). One key to immunity is having little proteins called antibodies produced by the immune system that specifically bind to foreign things (antigens) like viral proteins. Antibodies that bind to antigens on the surface of the virus can potentially block the virus from getting in (we call these neutralizing antibodies, or nAbs). The 2 main surface antigens for influenza are Hemagglutinin (HA), whose type determines half of the viral subtype (e.g. H1) and Neuraminidase (NA), whose type determines the other half (e.g. N1). Analogously, the main surface antigen for SARS-CoV-2 is Spike. 

Your body makes antigen-specific antibodies in response to an infection and vaccines try to trick your body into making antibodies without you getting sick – such as by injecting you with an inactivated version of the virus so your immune system learns to “recognize” and block it. So you want to make sure you show it the right “mug shot” during training.

One reason flu vaccines aren’t always effective is that there are multiple strains of flu, each with slightly different versions of the proteins which serve as antigens – epidemiologists try to predict what dominant strains will be circulating and put those in the vaccine, but they don’t always guess right, so a vaccine might train your body really well to recognize and stop one strain of the virus, but not the strain of the virus that you actually get confronted with. So you trained the immune system with a picture of Lex Luther and then then Darth Vader shows up. 

Alternatively, even if they chose the right strain, the strain might have evolved to “escape” antibodies. So you might get vaccinated against the correct strain, but then, by the time the virus finds you, it’s built up mutations that make its HA & NA look different, so those antibodies you made don’t “recognize” it. It’d be like Lex Luther grew hair or something.

This phenomenon, whereby a viral strain accumulates mutations over time which make it resistant to antibodies made against its ancestor is called “antigenic drift.” It happens a lot with the flu because influenza evolves rapidly, much faster than coronaviruses. Both viruses have an RNA genome. This means their genetic blueprint, which includes “recipes” for making their proteins, is written in RNA (really similar to DNA), and mutations (letter changes) in this RNA can therefore lead to small changes in the proteins. Influenza has a pretty sloppy RNA copier (RNA polymerase) which makes a lot of mistakes and, unlike coronaviruses, it can’t proofread. So mistakes (mutations) happen – most are neutral, some detrimental to the virus, but if they help the virus survive and infect more cells they’ll outcompete other viruses and gain dominance. Natural selection playing out on a teeny tiny scale. 

So, antigenic *drift* is gradual, usually 1 letter change at a time. But you can also have more dramatic, sudden changes in the antigenic protein through something called antigenic *shift.* This is where a cell is infected with multiple influenza viruses at the same time and they “swap” genes for their antigens. So, for example, if a cell gets infected with an H3N2 virus and an H1N1 virus, when the cell packages up the viruses to ship out, their genes can get mixed up (reassorted) and “oh sh*t you’ve got antigenic shift!” You might, for example, get an H3N1 or an H1N2. note: You can also have antigenic shift where the subtypes don’t change (e.g. H1N1 + H1N1 = different H1N1). 

Importantly, antigenic *shift* cannot happen for all viruses (and we don’t have to worry about it for SARS-CoV-2). It *can* happen for flu because influenza has its genome split up among 8 separate RNA strands (segments). The gene for HA is on segment 4 and the one for NA is on segment 6. So they can be swapped independently. And we definitely have to worry about this, because it can lead to the birth of pandemic strains, especially when influenza viruses that “grew up” in different animals come together. There are 18 HA subtypes & 11 NA subtypes, but they don’t all infect humans (knock on wood…) and H1N1 & H3N2 are currently the most prominent subtypes co-circulating among humans

so, to review:

antigenic drift

  • gradual accumulation of point mutations in HA & NA that let them escape antibodies developed against influenza HA & NA
  • unlike the coronavirus RdRP, which has a proofreader, the influenza RdRP is highly error-prone and those errors don’t get fixed. Most of these errors are neutral, or even harmful to the virus, but mutations that happen to be beneficial have a growth advantage. And if I virus has a mutation that prevents it from binding to antibodies someone has, it can sneak past the immune system

antigenic shift

  • reassortment of gene segments across different viral strains that are infecting the same host
  • this can lead to viruses with pandemic potential, as was shown with the 2009 H1N1 epidemic – that virus came about when parts from several H1N1 viruses swapped parts with each other, introducing people to antigens they didn’t have immunity to

quick note about what HA & NA actually are because you know it’s killing me not to tell you about these sugar scissors (NA) and cell-sticking glue (HA)! Both are surface glycoproteins. Influenza (similarly to the coronavirus) is an enveloped protein, so it travels as a membrane-bound sack of RNA and some proteins. NA & HA stick out from this viral membrane and are coated with sugars (glycans). Proteins, including receptors, on our cell surfaces are also coated with sugars, and HA finds one of these sugars, Sialic Acid (SA), especially sweet. HA binds to SA, allowing the virus to dock on (analogous to how the coronavirus Spike protein binds to the ACE2 receptor). After docking, the influenza virus gets “swallowed” by the cell in a process called endocytosis, which traps it in a “quarantine bubble” in the cell called an endosome. pH drop in the endosome causes HA to shape-shift (undergo a conformational change) which leads to fusion of the viral membrane with the endosome membrane, releasing the viral stuff into the cell where it goes to work hijacking the cell to make more viral copies. 

When those viral copies are ready to go infect other cells, they need to get out, which they do by budding from the cell surface. Problem is, they get stuck by SA on their way out. Thankfully (for them at least…) they have SA scissors in the form of NA. NA cuts the SA, allowing the new viral particle to be released from the cell surface. NA can also be helpful for freeing the virus from “decoys” like SA-coated mucin protein in your mucus. But the virus needs to make sure it balances its glue and scissors, so the surface has about a 4:1 ratio of HA:NA. “Fun fact:” Tamiflu is an NA inhibitor that mimics SA. 

Now that you’ve allowed me a brief geek-out, let’s get back to discussing vaccines (and then end with some more technical geek-out-ing).

Most influenza vaccines these days are quadravalent, meaning that they contain 4 strains. So they introduce your immune system to 4 different HAs and 4 different NAs. And note that “strain” is different from “subtype,” so you can have 4 different strains of H1N1s, for example. Most Influenza A strains currently circulating among humans are of the H1N1 & H3N2 subtypes. When you see H/N subtype stuff that’s usually talking about Influenza A. There are also 2 genetic lineages of influenza B, Yamagata and Victoria, which have different HA proteins, and circulate among humans. Quadravalent influenza vaccines contain 2 B viral strains (one from each lineage) and 2 influenza A strains (1 H1N1 & 1 H3N2). 

As we talked about, the point of a vaccine is to introduce you to a virus in a way it won’t hurt you. There are multiple ways to do this and the main types of flu vaccines are: inactivated (IIV)(totally dead virus), live attenuated virus (LAIV) ((nearly dead virus), and “recombinant” (viral proteins). 

Most of the vaccines are egg-based, but some are cell based. For these, they take a version of the virus that has adapted to grow well in eggs or cells and then they co-infect those eggs or cells with the strain they want to vaccinate against. Then they allow genetic reassortment (segment swapping) to happen. The cells churn out tons of different combos & what the manufacturers look for is a virus that has the HA & NA they want in the background of the egg- or cell-adapted strain. This way you get a lot of virus with the antigens you want. They isolate that winning combo & use it to infect more eggs or cells, get them to make a lot of virus, and then kill or weaken the virus with heat and/or chemicals. For the cell-based ones they actually then break apart the virus and purify and administer just the HA & NA. 

Commercial names for standard-dose inactivated flu vaccines include the one I got, Fluzone, as well as Afluria, Fluarix, FluLaval, & Fluarix. There are also a couple of “stronger” ones recommended for people 65 or older: Fluzone High-Dose, which as the name suggests, has a higher dose (4X the standard dose), and Fluad. Fluad has the same does as the standard ones, but it also has an added chemical called an ADjuvant which helps the immune system really recognize something foreign just got injected. The adjuvant it uses is a squalene oil-and-water mix called MF59. (squalene might sound scary but it’s a natural oil that our own skin makes).

Egg-based vaccines have some downsides. One is that some people have allergic reactions to egg proteins. Another is that sometimes the HA gets adapted to chicken cells, which don’t have one of the main types of sialic acid that HA likes). People with egg allergies can get Flucelvax, which is made in cell culture. note: Flucelvax is only trivalent,

Those are all inactivated vaccines, and they get injected into your muscle. There is also an egg-based live attenuated vaccine called FluMist which gets sprayed up your nose. 

You know how I said the flu virus evolves quickly? Well, the egg and cell-based production methods take a long time and sometimes the circulating virus antigenic-drifts during the production time so that the virus in the vaccine is too different from the circulating virus to be of much use. Additionally, as we saw with the sugar, the antigens might an adapt to grow well in chicken eggs or cell culture and those adaptations make it less like the original virus you want to introduce the person’s body to. 

To get around these problems, one vaccine, Flublok, just gives you the viral HA proteins. They take the strains of interest, isolate the HA gene and then stick that gene into a circular piece of DNA called a plasmid and stick that into expression cells to make the protein which they then purify and inject. They actually make them in insect cells using a baculovirus system, similarly to what I do in the lab to express my proteins!

Here’s a really good article explaining the pros and cons of different methods: https://cen.acs.org/pharmaceuticals/vaccines/Flu-shots-arent-always-effective/98/i1 

and some more great information from the CDC: https://www.cdc.gov/flu/vaccines-work/vaccineeffect.htm 

And remember – it’s not just about you when it comes to vaccination against the flu!

A couple of other, more detailed/technical notes about flu vs. coronavirus biology. Note: I’m going to use “the coronavirus” and SARS-CoV-2 interchangeably, but there are other coronaviruses including SARS-CoV (the original), MERS, and 4 common-cold-causing coronaviruses). Influenza is a virus in the Orthomyxoviridaefamily. Within this family there are 4 “genera:” A, B, C, & D. A & B are the main ones that cause problems for humans, and I’m going to focus most on Influenza A unless otherwise specified. Influenza is similar to SARS-CoV-2 (of the Coronaviridae family) in that both hold their genetic information in a single-stranded of RNA, but their genomic makeup is different in a couple of key ways. 

As I talked about above, the most obvious (and one of the most consequential) ways they differ is that, whereas coronaviruses squish all of their genetic info into 1 “long” strand of single-stranded RNA,  influenza’s genes are spread out among 6-8 separate strands of single-stranded RNA (referred to as segments) (influenza A has 8 segments). This is kinda analogous to chromosomes, except these segments are single-strands of RNA, whereas our chromosomes are double-stranded DNA. And, unlike our chromosomes, which each contain lots and lots of genes containing recipes for lots of different proteins, each viral RNA segment only has 1 or 2 genes. As we saw above, if a cell gets infected with 2 different influenza viruses at the same time, they can “swap segments” – a phenomenon called genetic reassortment – and this can lead to antigenic shift which can lead to pandemic strains. 

Influenza viruses also differ from coronaviruses in that the copy of RNA flu particles contain, and thus which enters an infected cell, is “negative-sense” – this means that the virus needs to make a complementary copy of the RNA before it can get the cell to make proteins based on the instructions it contains. Coronaviruses, on the other hand, are “positive-sense” – so they can get right to work making proteins once inside a cell. 

Basically, there are 4 RNA letters (nucleotides) and thanks to something called “base pairing” whereby each of the 4 RNA letters can specifically bind to one other letter (A to U and C to G), you can use one strand of RNA to make the complementary strand of the RNA – and then that complementary strand can be used to make an exact copy of the original strand – which can be used to make an exact copy of that complementary strand. But the 2 strands are not identical, and the protein-making instructions only “make sense” to the protein-making ribosomes in one of the strands – the “sense strand” or “positive strand.” The instructions are gibberish in the other strand, which we call the antisense strand or “negative strand”

genetic structure

  • influenza: 8 segments of single-stranded, negative-sense RNA
  • coronavirus: 1 strand of single-stranded, positive-sense RNA

docking

  • influenza: hemagglutinin (HA) binds to sialic acid (SA) on cell surface
  • coronavirus: Spike protein binds to ACE2 receptor on cell surface

membrane fusion

  • influenza: virus is “swallowed” by the cell (endocytosed) -> low pH in the endosome triggers fusion
  • coronavirus: protease cleavage at the plasma membrane or in an endosome triggers fusion

exit

  • influenza: gets trapped on cell surface because HA binds SA -> neuraminidase (NA) cuts it free
  • coronavirus: “no problems”

what’s most obvious to immune system (and thus most likely to be “antigenic” (have antibodies made against it)

  • influenza: HA & NA
  • coronavirus: Spike

you can find more on coronavirus-y topics here: https://bit.ly/covid19bbresources ⠀

and more explanations about all sorts of things: #365DaysOfScience All (with topics listed) 👉 http://bit.ly/2OllAB0 ⠀

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