The news these days is dominated by the one big story: the COVID-19 pandemic. Since the first reports of infection surfaced in China sometime in late 2019, the novel coronavirus that causes the disease, bloodlessly dubbed SARS-CoV-19, has swept around the globe destroying lives, livelihoods, and economies. Getting a handle on the disease has required drastic actions by governments and sacrifices by citizens as we try to slow the rate of infection

As with all infectious diseases, getting ahead of COVID-19 is a numbers game. To fight the spread of the virus, we need to know who has it, where they are, where they’ve been, and whom they’ve had contact with. If we are unable to gather the information needed to isolate potential carriers, all that we can do is impose mass quarantines and hope for the best. Hence the need for mass COVID-19 testing, and the understandable hue and cry about its slow pace and the limited availability of test kits.

But what exactly do these test kits contain? What makes mass testing so difficult to implement? As we shall see, COVID-19 testing is anything but simple, even if the underlying technology, PCR, is well-understood and readily available. A lot of the bottlenecks are, as usual, bureaucratic, but there are technical limits too. Luckily, there are clever ways around those restrictions, but understanding the basics of COVID-19 testing is the best place to start.

Going Hyperbolic

Currently, the only way to detect an active SARS-CoV-2 infection is by the use of polymerase chain reaction, or PCR. We’ve covered PCR in some detail before, but briefly, PCR is a laboratory method that relies on the cellular machinery that allows DNA to replicate itself. PCR is a three-step process:

1. Denaturation, which uses high temperatures to break the hydrogen bonds between base pairs in the DNA double helix, creating complementary single strands of DNA;
2. Annealing, which occurs as the temperature of the reaction is lowered and short oligonucleotide primers, specific for a section on the target DNA, bind to the single strands;
3. Extension, where the DNA between the primers is filled in by an enzymatic reaction. The result is a double-stranded DNA segment, which then goes through subsequent rounds of denaturation, annealing, and extension, resulting in exponential amplification of the specific target DNA.

If the DNA you’re looking for is present, the PCR process will amplify it to a large enough amount that it becomes easy to detect. If what you’re looking for is absent, it will not be amplified and it becomes equally easy to note its absence.

Molecular biology techniques in general, and PCR specifically, are incredibly powerful and flexible technologies. In the case of viruses like coronaviruses, that flexibility is advantageous because the viral genome is contained in a single-stranded molecule of RNA rather than DNA. Detecting the presence of the viral RNA in a patient sample, which is generally a throat swab containing viral particles expelled from the lungs and pharynx, requires first isolating the RNA, then turning it into DNA with the enzyme reverse transcriptase. That DNA is then amplified using PCR.

In either PCR or RT-PCR, detection of the amplified DNA region is accomplished by tagging the reaction with fluorescent probes. The probes bind nonspecifically to DNA, and when produced in abundance by PCR generate an easily detected signal. In actual practice, the kinetics of the PCR reaction are monitored by measuring the fluorescence after each cycle of PCR. This is known as quantitative PCR, or qPCR; when couple with reverse transcriptase, the process is called RT-qPCR.

PCR’s ability to detect the specific genetic signature of SARS-CoV-2 relies on the use of carefully selected primer oligonucleotides. Using the viral genome data published by the Chinese in January, the main primers were designed to amplify the genes coding for two viral encapsulation proteins. Along with those are primers for all SARS-like coronaviruses, as well as a primer for a human gene that should always be present, which acts as a positive control to make sure the reaction worked.

It’s In The Environment

Like any diagnostic or therapeutic technology, testing for COVID-19 is tightly controlled by regulatory agencies around the world. As frustrating as the delays caused by the necessary certifications may be, they’re understandable given that human lives are at stake. An emergency situation such as the current pandemic no doubt will relax some of those bureaucratic necessities, but they’re not likely to be eliminated entirely. Still, the simplicity of PCR and the availability of the reagents and instruments needed to perform the test are tempting targets for biohackers who are eager to do their part.

Fortunately, there’s a large unmet need for environmental testing that biohackers can pursue. Chai, a biotech company in Santa Clara, offers a low-cost, open qPCR instrument that’s capable of the exact kinds of protocols being used for human coronavirus testing. The instrument uses a Beagle Bone Black and while it is not cheap at $5,000 or so, it’s an order of magnitude less expensive than commercial, certified qPCR machines. Chai is currently marketing “environmental test kits” that can be used to swab doorknobs, desks, or other hard surfaces that might harbor droplets from an infected patient’s cough or touch. Thanks to PCR’s signal amplification ability, the samples can contain as little a single copy of the viral genome and still be detected.

Chai is clearly not playing games here and specifically caution against use with human samples. They are strictly in the environmental testing market, but that’s a good thing. Coronavirus can only get into the environment once it escapes from a host, and the virions can only survive for a limited amount of time away from a host cell. That makes environmental testing an excellent proxy for the viral load of the humans in that environment, and can be used to get an idea of the dynamics of that load over time. By making relatively cheap qPCR tests possible, Chai is offering a backdoor solution to the problem of limited human testing.

It’s unlikely that human COVID-19 testing via RT-qPCR can be made to go much faster than it already is. Even when the logistics problems regarding the manufacture and distribution of test kits are solved, there are only so many certified qPCR machines to go around, and only so many technicians to run them. Add to that the likelihood that the techs themselves will get sick, and human testing can only proceed so fast. Innovations like drive-through testing are only intended to protect health care workers and the testing environment (since the throat swab often causes patients to cough, drive-through testing where the patient only cracks the car window means a cough doesn’t contaminate an exam room). And such testing will only increase the number of samples that need to be processed.

Despite these challenges, the availability of PCR as a diagnostic method has been a boon to epidemiology in general and the fight against COVID-19 specifically. And if others follow Chai’s lead in environmental testing, with any luck we may soon see a flood of new data that will reveal the true scope of this pandemic and finally let us start making some headway against it.