Monthly Archives: April 2020

COVID-19; Immunology and the Infection Cycle, 4/22/20

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Sorry for the delay, but this one has taken a lot of time and thought to put together (and reading some 80 odd Covid research papers). My goal in this edition of Covid Science Theater 2020 is to talk about what happens when the virus enters our body, infects our cells and subsequently leads to either mild disease or more severe infections. It’s going to be a fairly dense article, but I’ll do my best to keep the science and terminology to something generally understandable and hopefully educational.

For those who don’t want to delve too much into the specifics of all the virology, immunology and pathology I’ll provide a short 1 paragraph set of clifs notes here. The virus most commonly enters the host through mucus membranes (eyes, nose, mouth) and infects vascular endothelial cells and cells of the lungs, kidneys, GI tract and begins to replicate. The body initially responds to the virus through a host of Innate Immune mechanisms; these generic counter measures are deployed against all invading pathogens as a first line defense and are not specific to the invading pathogen. Unfortunately, these initial immune responses aren’t always adequate to contain the virus (the virus sometimes evades destruction, other times the virus just overpowers the immune response) so our body deploys a second type of response known as the Adaptive Immune response. In this phase, T-cells and B-cells are primed to respond to the specific infectious agent (here, SARS-CoV-2). Often this two-pronged approach works to contain the infection, eliminate the virus and build up lasting memory to subsequent infections. Unfortunately in some people the virus spreads too rapidly and the immune response doesn’t respond appropriately, leading to destruction of their organs (notably the lungs) and potentially death. Sometimes this more severe outcome is caused by the virus itself, but more often it seems to be caused by an overzealous immune system trying to play catch-up. So there’s the quick and dirty; in the following paragraphs I’ll go into more detail about Viral Entry/Binding/Replication, Early Cellular Responses, Clinical Symptoms, Adaptive Immune Response and What Happens and Why the Immune System Sometimes Fails.

The information in the following paragraphs comes from a combination of basic immunology principles (Kuby Immunology textbook), observations and early research released about Covid-19 and conclusions drawn from earlier studies of SARS-CoV-1 (a virus that is very similar to the current SARS-CoV-2, but with some caveats of course). As Covid-19 is still a new disease, we are constantly learning new things about the virus, infection cycle and pathology, so while what I outline here is based on a lot of research, there are definitely aspects of this virus that we don’t fully understand, and need further investigation.

Graphic of the structure of a SARS virus, the S, M and E proteins are the most important in regards to host recognition, Li et al 2020.

Viral Entry and Replication

Coronaviruses get their name from the hallmark shape, a circular capsid (or shell) that is spiked with proteins on the outside and sheltering the virus genetic sequence on the inside. The Spike (S), Membrane (M) and Envelope (E) proteins make up the majority of the viruses outer shell, while the Nucleocapsid (N) protein found inside the virus assists in viral replication. This small assortment of proteins, plus a few others, make up the bulk of the very simple viral structure (Weiss 2005, Li 2020). The novel coronavirus 2019 (COVID-19) shares a lot of homology or similarity with the original SARS virus that was discovered in 2002, genetically 80% similar, while being 76-95% similar for the major proteins listed above (Xu 2020). This allows researchers to draw a lot of conclusions from previous research on SARS-CoV-1, though we must be careful when doing so, as there are some known (and unknown) differences between the two viruses. The infection cycle starts with the virus gaining entry to the host, usually through mucus membranes of the eyes, nose and mouth. Once inside the virus often begins it’s infectious cycle by infecting vascular endothelial cells that line vessels throughout the body. While different viruses have different mechanisms by which they enter host cells, SARS-CoV-2 binds to the ACE2 receptor using its spike protein (same as SARS-CoV-1), allowing it to enter the host cell (Jia 2005, Walls 2020). Like most viruses, SARS-CoV-2 then goes through a multi-stage process by which it hijacks some of machinery inside our own cells to in order to replicate, escape and subsequently infect more cells in a continual cycle (Frieman 2008).

Overview of the SARS viral life cycle inside the host, Frieman et al 2008.

Early Cellular Response

Thankfully our body has a whole host of immune mechanisms it utilizes to deal with infectious agents of all types. Almost as soon as an invading pathogen has infected our cells the immune system starts going to work. The Innate Immune response is our constantly active sentinel, whose cells are constantly circulating all over our body just looking for foreign invaders to attack and kill. These innate cells use Pathogen Associated Molecular Patterns (PAMPs), or markers of foreign invaders, as the initial signals something is wrong and that it’s time to go to work (Li 2020). Some cells go to work directly attacking the virus and infected cells in an attempt to destroy the virus, others release signaling molecules known as cytokines and chemokines that recruit other cells to help in the fight (Frieman 2008), and some cells just go ahead and sacrifice themselves in an effort to prevent the virus from hijacking them, a process known as apoptosis (Lim 2016).

Symptoms: What and How They Manifest

While this system works well for many invading pathogens (why we are not sick all the time), allowing our body to control the infection, many viruses (and bacteria) have evolved mechanisms by which to evade, subvert and co-opt the immune response to their advantage. For SARS-CoV-2 it seems to be able to prevent the host immune system from activating one of it’s key anti-viral signaling pathways, the Type 1 Interferon pathway (Lim 2016, Li 2020, Frieman 2008). While it is not known exactly how the virus subverts this system a few hypotheses involve the Nucleocapsid protein (Lim 2016), other non-structural proteins (Lim 2016), and some of the SARS enzymes (Chen 2014). So by reducing the host immune response the virus is able to more effectively replicate and spread, leading to a more systemic infection. This is when we start to experience more of the hallmark symptoms of the infection; fever, sore throat, coughing, fatigue, pulmonary inflammation leading to shortness of breath and possible pneumonia and lymphopenia (a decrease in lymphocytes, more on that later) (Huang 2020, Zhu 2020). Most of these symptoms are a physical outcome of the body’s ongoing fight with the virus, trying to delicately balance destroying the invader, while preserving the host organs and system. The fever is the immune system’s attempt to raise the core temperature enough to burn out the infection. The sore throat/cough is an outcome of our immune system attacking infected cells of the airways and trying to expel the invader (mmm mucus), same for the pulmonary issues (initially, more on this later too). While many of these symptoms may be scary and uncomfortable they are often a normal part of our body’s healing process when dealing with a foreign invader. So under normal circumstances, it’s best to rest and let your body do it’s thing, unfortunately this doesn’t always go as planned, as we’ll find out in the following sections….

Adaptive Immune Response; Stage 2

In the previous two sections you’ve seen how our well intentioned Innate Immune system can sometimes fail leading to illness, thankfully the body has a backup, the Adaptive Immune response. This secondary wave of the immune response goes into action very soon after the initial infection (several hours to few days, infection dependent) and is mostly comprised of two cell types; T-cells and B-cells. When the levels of virus in the body start to rise, several of the innate immune cells can act as activators of the adaptive immune response, taking pieces of the virus to specialized activation centers know as lymphoid organs. These centers of immune activation are spread all over our body and are the primary site of pathogen specific antigen (virus pieces) presentation. The antigen presenting cells (Dendritic cells are most prominent) present the virus to the T-cells and B-cells as if locks in a door, allowing the T-cells and B-cells to go to work making specific keys (receptors and antibodies) that can attack and destroy the pathogen in a very focused manner. The outer proteins that make up the viral capid (proteins S, M, E) tend to be the most effective as this is what is visible to our body when intact virion are released (Liu 2017). So the body makes a whole army of these specific cells that traffic to the sites of infection; T-cells directly attack the virus and infected cells, while B-cells make antibodies that bind to parts of the virus, preventing them from entering new cells and marking them for destruction (Liu 2017).

These two arms of the Adaptive Immune response are also what comprise our immunological memory. Virus specific T-cells and antibody producing B-cells remain dormant in specialized lymphoid organs (sometimes they also remain in circulation), just waiting for the virus to turn up a second time. This time since they are already primed and ready to go, memory T-cells and B-cells start attacking the virus almost immediately, usually preventing the virus from spreading and preventing us from getting sick. Studies of SARS-CoV-1 have found both memory T-cells and memory B-cells (producing neutralizing antibodies) that are capable of rapidly responding to viral reinfection (Li 2020, Liu 2017, Channappanavar 2014). In human patients who recovered from SARS-CoV-1 infection anti-SARS antibodies and memory T-cells were found in most patients up to 24 months after infection (Liu 2006, Ka fai 2008, Liu 2017). While antibody responses did decline over time in SARS-CoV-1 patients (many undetectable at 6 years), memory T-cell responses were conserved for up to 11 years after infection (Tang 2011, Ng 2016, Liu 2017). Similar high quality neutralizing antibodies have been found in COVID-19 patients, but since the disease is so new the longevity of memory responses to this new virus aren’t exactly known. Encouragingly, since SARS-CoV-2 is so similar to the original SARS virus, and lab testing has even shown that their be might cross-reactive protection between the two diseases (Walls 2020), there is much hope that the long lasting memory responses seen for SARS-CoV-1 would also apply to those who have recovered from COVID-19. All of this evidence, both old and new, does inspire a lot of hope that a functional vaccine would both be likely and very effective in providing some duration of immunity from COVID-19, but how long remains to be seen.

Graphical overview of the many cells and pathways involved in the host immune response to SARS. It’s a complex set of feedback loops and interactions with a lot of variables. Li et al 2020.

When the Immune System Fails, Severe Disease

The reason COVID-19 is such a scary disease, isn’t because our immune system has no problem fighting it off, but because in some percentage of the cases (uncertain, but estimates are as high as 10-20%) patients need to be hospitalized due to severe complications. If our immune system is so complex and so strong, why do patients with COVID-19 get so sick that they need hospital care? It comes down to numerous very subtle things this virus does that are different than coronaviruses that cause the common cold. One is the effect SARS-CoV-2 has on Type 1 Interferons mentioned earlier, reducing the body’s initial response to infection. Another early symptom seen in many severe cases is lymphopenia, or a loss of lymphocytes (notably T-cells) early on in disease (Huang 2020, Schmidt 2005, Weiss 2005). While the exact cause of this loss of T-cells is not known, it is hypothesized that the viral proteins may lead directly to T-cell death as a mechanism of immune evasion (Lim 2016, Li 2020). These mechanisms of avoiding immune detection along with the efficiency of viral replication can lead to an out of control infection very quickly.

But in the end, it’s only partially about the virus, and largely about an overexuberate immune response. In an attempt to catch-up to the wide-spread infection the immune response goes into overdrive, ramping up a lot of the inflammatory cells and signaling molecules that tell the body to attack the infection (Li 2020). This response does in fact kill the infected cells, but it also destroys lung tissue (primary target), vascular tissue, liver tissue and other infected tissues (Tian 2020, Schmidt 2005). This is often when the more obvious signs of pneumonia set in; the lungs fill with fluid, the efficiency of aveoli decreases (oxygen absorption) and breathing becomes very labored and difficult. This is the tricky thing about COVID-19, making our immune response more efficient would help prevent early infection, but later on would lead to increased tissue damage. But if we reduce the immune function of the body to prevent self-inflicted tissue destruction, we run the risk of allowing the virus to run rampant throughout our body. COVID-19 is a tricky disease to treat for these reasons, and because the disease severity has a wide range of outcomes for different people. In some, infection is very mild and asymptomatic, in others, their entire body shuts down as the virus (and immune system) destroys the host from the inside. The reason many comorbidities are important as risk factors for severe disease is that most of them either affect the immune system or lung function. Obesity, diabetes, auto-immune diseases all alter the immune system’s ability to function, making it harder to fight off the virus. COPD and asthma (though less prominent then thought) make the host pulmonary system more sensitive to damage caused by the virus and immune system.

But not all hope is lost! Because of the large body of evidence suggesting that SARS viruses create robust lasting immunity, this means a vaccine might be very effective at protecting most of the population. Also, now that there are many patients who have recovered from COVID-19, tests are underway to examine if using their plasma (containing antibodies) can help patients who are suffering from more severe cases of the disease (works for other viruses like Ebola). We also have several promising anti-viral agents that are already in clinical trials being tested against COVID-19, with hopes that one or more of them will help improve patient outcomes and be ready for use later this year. Unfortunately all of this does take time, meaning we won’t have a cure next month, but by slowing the spread of the virus, not only do we allow hospitals to manage the patient load, but we allow all the scientist out there to catch-up and produce much needed data, therapies and vaccines.

Thanks for reading. If you see any mistakes please bring them to my attention and I will correct them ASAP. If you have additional questions or want to discuss the immune response in more detail (this is a very high level overview) I’d be happy to do so via text or email. Stay safe and stay healthy.

Literature Citations:
Chan et al, Serological Responses in Patients with Severe Acute Respiratory Syndrome Coronavirus Infection and Cross Reactivity with Human Coronaviruses 229E, OC43, NL63. Nov 2005, Clinical and Diagnostic Laboratory Immunology.
Channappanavar et al, T cell-mediated immune response to respiratory coronaviruses. May 2014, Immunology Res.
Chen et al, SARS coronavirus papain-like protease inhibits the type 1 interferon signalling pathway through interaction with the STING-TRAF-3 TBK1 complex. Jan 2014, Protein Cell.
Frieman et al, SARS Coronavirus and innate immunity. 2008, Virus Research.
Huang et al, Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China. Jan 2020, Lancet.
Jia et al, ACE2 Receptor Expression and Severe Acute Respiratory Syndrome Coronavirus Infection Depend on Different Human Airway Epithelia. Dec 2005, Journal of Virology.
Ka-fai Li et al. T cell responses to Whole SARS Coronavirus in Humans. Oct 2008, Journal Immunology.
Li et al. Coronavirus infections and immune responses. Jan 2020, Journal of Medical Virology.
Lim et al. Human Coronaviruses: A Review of Virus-Host Interactions. 2016, Diseases.
Lu et al. Immune responses against severe acute respiratory syndrome coronavirus induced by virus-like particles in mice. June 2007, Immunology.
Ng et al, Memory T cell responses targeting the SARS Coronavirus persist up to 11 years post-infection. March 2016, Vaccine.
Schmidt et al. Coronaviruses with a special emphasis on First Insights Concerning SARS. 2005, Birkhauser Advances in Infectious Diseases.
Tang et al, Lack of Peripheral Memory B cell responses in Recovered Patients with Severe Acute Respiratory Syndrome: A Six-year Follow-up Study. May 2011, Journal of Immunology.
Tian et al, Pulmonary Pathology of Early-Phase 2019 Novel Coronavirus (COVID-19) Pneumonia in Two Patients with Lung Cancer. Feb 2020, Journal of Thoracic Oncology.
Walls et al, Structure, Function and Antigenicity of SARS-CoV-2 Spike Glycoprotein. Apr 2020, Cell.
Weiss et al, Coronavirus Pathogensis and the Emerging Pathogen Severe Acute Respiratory Syndrome Coronavirus. Dec 2005, Microbiology and Molecular Biology Reviews.
Xu et al, Systematic Comparison of Two Animal-to-Human Transmitted Human Coronaviruses: SARS-CoV-2 and SARS-CoV. Feb 2020, Viruses.
Zhou et al, Clinical course and risk factors for mortality of adult inpatients with COVID-19 in Wuhan, China: a retrospective cohort study. March 2020, Lancet.

COVID-19 and Masks, 4/5/20

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There’s been a lot of debate and misinformation floating around about the use of masks for the general public as a measure to prevent the spread of infectious diseases, specifically Covid-19 in the United States. Do they help, do they not? Is an N95 really better than a surgical mask, is this better than a cloth mask? How and when should they be used? On 4/3/20 the CDC in the United States finally came out with a blanket recommendation that ALL citizens of the United States wear some sort of face covering whenever in public. This was a dramatic change of direction from the previous recommendation that masks were completely unnecessary, except for front line hospital workers and for the infected. In this rendition of Eric’s Science Corner I’ll do my best to present some of the data and studies that have looked at the questions above, in an attempt to clarify the misunderstandings and the mixed messages. The topics I’ll try and cover are; what are the different types of masks and what are they designed to do? How useful are the different types of masks for the general public? And finally, a few best practices on how to wear and use a mask or face covering. Rule #1, just ignore anything Donald Trump says, now on with the info.

Defining Mask Types

To start there are three main categories of face masks that I’ll be discussing; fitted N95 respirators, professional grade surgical masks and cloth masks (variety of materials). There are numerous sub-categories for each and also other types of protective face wear I won’t discuss because they aren’t really relevant to the general population, only to those in the hospitals and those of us who work in laboratories. The first type is the fitted N95 respirator, these are face fitted respirator masks that have been certified to filter out approximately 95% of aerosols and particulate matter (when worn properly). You breath through either a small filtration unit in the front of the mask or directly through the filtering material of the mask, NOT around the sides (as it should be sealed). The professional grade surgical masks that many of us have seen in the hospitals are loose fitting non-sealed masks that are designed to block the wearer from inhaling large droplets/splashes and to block their respiratory emissions (protecting others around them). They are not designed to prevent the wearer from inhaling aerosolized particles as they are not sealed around the edges (cdc.gov, crosstex.com). The final group are the cloth masks which can be made from various materials. Their main purpose is to allow more comfortable widespread facial covering for the general public; to reduce the inhalation of larger droplets and to reduce one’s own exhalation and aerosol creation. These types of masks are not specifically certified in any way, though I will discuss the research that has been done looking at filtration, efficacy and utility of the different materials.

On the left is a standard N95 Respirator mask, on the right is a surgical mask.

So now on to how well do these different types filter out microparticles, specifically in regards to viral transmission (because that’s what’s on everyone’s mind). Numerous studies that compare N95s and surgical masks and how they prevent infection in hospital settings have shown both to be similarly effective when dealing with droplet based respiratory viruses like influenza (Randanovich 2016, Smith 2016). Laboratory testing of these two types of masks does confirm that the smaller the particle size, the better an N95 performs compared to a surgical mask (van der Sande 2008, Shakya 2016), thus they are more effective for those dealing with high level risk of aerosolized viral exposure. These two types of masks are certified, so it’s no surprise they perform fairly well, but what about the cloth and homemade masks? The first thing to consider is the type and thickness of material being use for the mask. Things that allow easy breathing or light to penetrate aren’t going to filter the air as efficiently, but if it’s too thick that you can’t breathe through it then it becomes extremely hot, uncomfortable and unwearable (and you breath around the sides, rather than through the material). One study comparing the filtration efficiency (of masks in a lab test, not on a person) of different materials found that items such as tea towels and cotton mixed fabrics did the best job of filtering particulate matter (up to 70% mean filtration) out of the air, while silk, scarves (like buffs), pillow cases and normal cotton T-shirts did not perform as well (45-60% mean filtration efficiency), with surgical masks being their standard (90-96% mean filtration)  (Davies 2013). When commercially available cloth masks were compared to surgical masks on humans (again in a lab) filtration efficiency was more variable; with cloth filtering out 30-50% of microparticles while surgical masks filtering out 60-90% of microparticles and N95s consistently filtering out 80-95% (Shakya 2016). The efficiency of filtration directly correlated to the size of the particle, with cloth masks performing the poorest on particles small than 1µm in size. So that’s a little background about how the masks are INTENDED to be used and how they function in a laboratory, how about in real life?

Use in the General Public

By now you’ve probably heard many times that the public should not hoard or use N95s because we need them for our frontline workers (very true) and they don’t work for the public (partially true). The first piece is that because of the size of this pandemic we don’t have sufficient supplies of N95s for highly trained hospital workers who are coming into direct contact with the virus on a daily basis, thus need this heightened level of protection, first (and most important) reason not to stock up or hoard them. The second is that for an N95 to be at it’s most useful and functional you have to have it fit tested, you need to be trained in proper techniques to don/doff a mask and you have to actually use it correctly (you can’t be taking it off to talk, to eat, to drink, basically you can’t break the seal unless in a clean contained environment). They are also designed to be disposable, meaning you can’t wash them, though sadly our healthcare workers are being forced into extreme measures to try and sterilize/reuse them for lack of options. For the general public a surgical mask would be a descent option because they are designed to reduce droplet transmissions and to block one’s exhalations (protecting those around you), but sadly our hospitals are also short on these too, so for now they need to be saved for the frontline works (and patients) where they’ll do the most good. Also remember that both of these are designed to be disposable, so can’t be washed and aren’t designed to be reused for weeks on end (like the public would need).

So this brings us to cloth masks and their use in the general public. Mistakenly the US government (CDC) originally came out saying that cloth masks don’t work and that they aren’t necessary. By now most people have realized this isn’t exactly true, because why else would they change their minds and recommend people wear them? Yes, cloth masks are NOT designed to stop all tiny viral particles (and aerosols) from passing through, and yes they are not highly efficacious, but that doesn’t mean they don’t help. While a cloth mask won’t fully stop one from inhaling aerosols and microparticles, they do filter out some of the smaller aerosols (30nm-1µm) but more importantly block larger droplet transmission both inward and outward (Davies 2013, Shakya 2016). So while they do filter some of the air you’re inhaling, the major benefit of a mask is to protect those around you by minimizing the amount of aerosols you create. This is especially true with the knowledge that those infected with COVID-19 can be asymptomatic but still capable of spreading the infection. For masks to be most beneficial we all should wear them in any public setting where we’ll be interacting with others (even if we’re socially distancing).

Best Practices for Masks

Now on to a few personal suggestions for best practices when using a face mask. Note that much of this stems from my own personal training having worked in Biosafety Level 3 laboratories (blood and aerosol transmitted infectious diseases) and in hospitals, but some additional guidance can be found on the CDC website (CDC.gov). First off, once you’ve made/acquired your mask, put it on at home and work on the fit, comfort, breathability. A mask that doesn’t stay on or that you can’t semi-comfortably wear (to the point you’ll touch it a lot or take it off) isn’t very useful. Look to make sure it fully covers your nose and mouth, has a pretty good fit around the bridge of your nose and the sides, and that it won’t slip down when you turn/move your head.

Once you’ve established it works, wear it around for 20-30min inside your house to get used to the idea of breathing through a mask. It’s probably going to be a bit awkward at first, as for most people they’ve never had to do it before. This exercise will make it easier to wear in public without thinking about it too much. Now on to that more critical step, wearing it out. The main times the mask should be worn is whenever you’re going into a public area where you might have close contact with others. If you’re just sitting in your car and driving around, no need to wear a mask, but if you go to the grocery store, pharmacy, liquor store, gas station, work, or even walk around your neighborhood it’s best to wear the mask to protect those around you, even if you don’t think you’re sick.

To put on the mask, do so BEFORE entering that public space, meaning your home entry if you’re walking around the neighborhood or inside your car before you walk into a shop. Then clean your hands off so that you are less likely to contaminate other surfaces (hand sanitizer or washing). When you’re wearing you mask you SHOULD NOT be taking it off or moving it off your nose/mouth until you’re back in your non-public safe area. Wearing it half the time, pulling it down half the time, taking a break to eat or drink in public negates some of the benefits and protection and also adds to the chance that anything you pickup on your hands will be transferred to your face. When you’ve exited the public space, wash/clean your hands then grab the strings/band of the mask and remove it (do not grab the front of the mask itself). If you have a washable reusable mask proceed to wash it with soap and water. Disposable masks are supposed to be discarded into the trash (hence why not ideal for daily use in public). While your mask is your barrier of protection, remember it’s not foolproof, and is merely a way to further reduce your risk of becoming infected and infecting others. IT DOES NOT change the fact we should be social distancing and providing each other space or that we should be staying at/near home and avoiding any unnecessary travel/errands. Wearing a mask is just another tool in our arsenal to help slow the spread of the virus and reduce transmission rates.

One last note about gloves. Wearing gloves for most people in a public setting is useless (yes I said useless). Gloves are a very effective piece of PPE for trained healthcare and lab workers, but in our daily lives most people treat gloves just like their normal hands. They touch common surfaces, pick up food items, open doors, text on their cell phone, touch their mask, etc. All of these practices together make the use of gloves just as bad as dirty naked hands. You’re better off just considering your hands as dirty whenever you’re in public and not touching any of your personal belongings (including that cell phone) until you’ve cleaned them. If you have to touch your phone or food items while in public, there are many ways to also clean these surfaces as well. Don’t waste gloves and don’t touch your face.

Citations
Balazy et al. Do N95s respirators provide 95% protection level against airborne viruses, and how adequate are surgical masks. American Journal of Infectious Control, 2006.
cdc.gov/hai/pdfs/ppe/ppeslides6-29-04.pdf . CDC Guidelines for Selection of PPE in Healthcare.
cdc.gov/niosh/npptl/pdfs/UnderstandDifferenceInfographic-508.pdf . Understanding the Differences, Surgical Masks, N95 Repsirators.
crosstex.com/sites/default/files/public/educational-resources/products-literature/guide20to20face20mask20selection20and20use20-202017.pdf . Guide to Face Mask Selection.
Davies et al. Testing the Efficacy of Homemade Masks: Would They Protect in an Influenza Pandemic. Disaster Medicine and Public Health Awareness, 2013.
osha.gov/Publications/osha3079.pdf . OSHA Respiratory Protection Guidelines.
Randanonvich et al. N95 Respirators vs Surgical Masks for Preventing Influenza amount Healthcare Personnel. JAMA, 2019.
Sande et al. Professional and Home-Made Face Masks Reduce Exposure to Respiratory Infections among the General Population. PLOS One, 2008.
Shakya et al. Evaluating the Efficacy of Facemasks in Reducing Particulate Matter Exposure. Journal of Exposure Science and Environmental Epidemiology, 2016.
Smith et al. Effectiveness of N95 Respirators vs Surgical Masks in protecting healthcare workers from acute respiratory infection: a systematic review and meta analysis. CMAJ, 2016.