Towards more efficient diagnosticsMay 10, 2020 0 By FM
While the rest of the world was completely unprepared to face the COVID-19 outbreak, three countries — Taiwan, Hong Kong and Singapore — used their earlier experience with disease outbreaks to reduce the spread of the virus. This suggests that it is possible to effectively contain the pandemic, even if you are situated close to China. These three countries have rapidly deployed testing, combined with digital surveillance, to trace infected individuals and have put in place strict quarantine for suspected cases. These measures, along with the build-up of large stockpiles of personal protective equipment, allowed them to fight back quickly. The majority of other countries reached the stage of community transmission before adequate testing was in place, including decentralized, point-of-care testing.
There are two major technologies at present for diagnosis of COVID-19, real-time polymerase chain reaction (RT-PCR) and antibody detection systems. The RT-PCR system is very sensitive and can detect even very low amounts of virus from inside of the mouth or nose. The first RT-PCR tests for COVID-19 were designed and distributed in January 2020 by World Health Organization (WHO). However, the test protocol is very complex, involving sample collection, shipping to a central laboratory, RNA isolation and reverse transcription followed by quantitative PCR. This protocol needs high-end laboratory infrastructure and logistics and has a high turnaround time of minimum of 1 to 2 days at best.
A similar test based on isothermal amplification-based technologies is under development for COVID-19 diagnostics. The isothermal techniques include recombinase polymerase amplification, helicase-dependent amplification and loop-mediated isothermal amplification (LAMP). Reverse transcription LAMP (RT-LAMP) uses DNA polymerase and four or six primers that bind at distinct regions in the target sequence of COVID-19, making it a highly specific test. In this assay, the patient sample is added to the tube and the amplified DNA is detected by turbidity, color or fluorescence. The challenge with LAMP is that it requires a long time for optimizing primers and reaction conditions.
Limitations of immunoassays
Immunoassays, on the other hand, can provide information about viral exposure and diagnostic evidence. Such serological assay systems detect immunoglobulin G and M (IgG and IgM) from the serum of COVID-19 patients. The antibody test needs to be run on a sample of blood from a patient and is not looking for the virus directly, but at the body’s capacity to produce antibodies against the virus. Not all antibodies are the same or created at the same time during the infection. The antibody IgM is the body’s first response to an infection and is normally produced within 5-10 days after the infection takes hold, peaking at 21 days post-infection. That time frame is very important, because if an individual has just developed symptoms for coronavirus, it will take approximately a week for a person’s body to raise IgM antibodies. Even though, with coronavirus, we have evidence of IgM being present in the blood within 1 day of symptoms, that isn’t going to be a reliable test at that stage as there likely won’t be large amounts of IgM to detect. IgG shows that someone has had the virus and is now protected from it. This can be detected in a patient’s blood around 10-14 days after infection. These limitations make immunoassays less reliable and sensitive. Recently, Indian Council for Medical Research (ICMR) has given a directive to state governments that antibody tests cannot replace RT-PCR testing.
Integration of decentralised testing
Neither RT-PCR nor immunoassays are going to be solutions to combat this pandemic. However, the latter is faster and delivers results within 20-60 minutes. At a later stage, flow immunoassays can be mass manufactured and can be used by trained personnel. In case of this virus, the spike protein (S) facilitates entry of the pathogen into airway epithelial cells by binding to the angiotensin-converting enzyme 2 (ACE2) expressed on these cells.
Companies like Neuome Technologies Pvt. Ltd., in collaboration with Institute for Applied Research and Innovation (India), and GE Life Sciences, in collaboration with Halifax and Sona Nanotech from the USA, are focusing on developing lateral flow assay systems to detect the virus by exploiting the above mechanism. These technologies will be able to directly measure actual viral particles in concentrations as low as 30 particles, in as little as 5-10 minutes, and can be used on-the-spot. They will be similar to a pregnancy strip test, but will use a buccal swab or saliva for detection. This technology will help in identifying even asymptomatic individuals in the early stages of infections. As it does not need any instrument, it can be deployed even in remote areas, at ports of entry (such as airports), for soldiers, social gatherings, malls and other places. Such integration of decentralised testing with epidemiological surveillance would go a long way in enabling public health authorities to reduce the spread of the infection.