** Slow place in Dhaka’s canals demarcation not acceptable ** Powerful earthquakes jolt Turkey and Syria ** Apparel sector eyes $15b Indian raw material support ** Belgium's Queen Mathilde Marie Christine visits corporate building, production house, day-care centre and different parts of knit composite manufacturing company Fakir Apparels Limited in Narayanganj district on Monday. PID photo ** President and PM express shock over quake in Turkey, Syria ** DB's final report clears Bushra, settles on suicide ** Turkey earthquake is country's worst disaster in 84 years: Erdogan ** IMF’s loan condition Additional Tk 65,000cr to be collected next year ** Major earthquake in Turkey, Syria kills nearly 2,300 people ** Bangladesh not sure to what end Indo-Bangladesh friendship is leading under Mr Modi's anti-Muslim stance ** PATHETIC INDEED! Geeta Biswas, a poor woman, failed to purchase OMS (open market sale) rice after standing in a long queue for hours. At last, she climbed into the truck for rice. When she tried to pick up the scattered rice in bags from the truck, the sellers took away the bags from her. Later she picked and took about one and a half kilos of rice. This photo was taken from Agrabad Government Commerce College area in Chattogram on Sunday. Agency photo ** Sending migrant workers BD-Malaysia agree to review MoU signed in 2021 ** Draft data protection law is a matter of concern: US envoy ** Pakistani former President Pervez Musharraf dies ** GDP growth in FY22 reaches 7.10pc ** US shoots down Chinese spy balloon ** The construction of a 300 feet wide and 12.5 kilometres long expressway from Kuril flyover to Kanchan Bridge to connect Purbachal New Town with Dhaka city is in the final stage. This photo was taken on Sunday. ** Bangladesh opens doors for 2,970 more Rohingya people ** Take necessary measures to stop theft of RMG goods on highway: BGMEA ** Influentials hamper reform process : Economists ** BNP announces march prog on February 11 ** Probe alleged enforced disappearances ** Stolen device worth Tk 47 lakh recovered ** Spy balloon row: High-altitude spying marks new low for US-China ties ** 642 killed in road accidents in January: JKS **

Saliva Tests: How They Work and What They Bring to Covid-19

11 July 2020
Saliva Tests: How They Work and What They Bring to Covid-19

Amanda Heidt :
Faced with an ongoing lack of protective equipment and testing supplies, medical professionals have been seeking alternatives to accurately diagnose cases of Covid-19, a pandemic that has caused more than 11 million cases and more than 530,000 deaths worldwide. Supplies of nasopharyngeal swabs were some of the first testing materials to run low in mid-March, prompting a pivot to nasal swabs. More recently, saliva-based testing has come forward as an attractive, low-cost alternative.
The first spit tests are already being sold to consumers, with more poised to apply for emergency use authorization from the US Food and Drug Administration soon. While saliva can be a crude sample for diagnosing disease using traditional PCR, it pairs well with a cheap PCR alternative known as loop-mediated isothermal amplification (LAMP), previously used to detect outbreaks of Zika and Ebola in resource-poor countries. Propelled by a global pandemic, researchers in the US and the UK are now modifying LAMP and assessing its utility as a diagnostic tool for Covid-19.
"By diversifying the possible choice of assay, you diversify the supply chain as well," says Robert Meagher, a chemical engineer at the Sandia National Laboratory who develops tools for diagnosing emerging diseases. Making additional testing options available to healthcare workers will help mitigate backlogs if one reagent or material runs low.
Saliva-based testing also offers an improvement over the standard nasopharyngeal swab because people can collect their own samples with minimal discomfort-simply spit into a sterile tube and mail it to a lab for processing. And due to an emergency use authorization given to Rutgers University in May, some tests can now be carried out by patients in their own homes, allowing personnel and protective equipment to be saved for when they are most needed. Andrew Brooks, the director of technology development at RUCDR Infinite Biologics, the Rutgers-affiliated biorepository that developed the test, says the use of in-home testing "completely mitigates the risk of contracting the disease while you're getting a test," and requires only gloves.
Several companies are selling Rutgers's kits directly to consumers through online orders, although all tests must be sent back to its lab in New Jersey for processing. Positive results are reported to local health officials, and individuals are advised on how to respond.
Universities and clinicians alike are now planning to incorporate saliva-based sampling into their workflow for diagnosing Covid-19. The UK government recently partnered with the molecular diagnostics company Optigene to develop a pilot study involving more than 14,000 people to test the efficacy of its saliva test. In the US, the University of Chicago will use spit tests to clear patients prior to elective surgeries in its hospital for the next several years, Evgeny Izumchenko, a UChicago oncologist who helped develop the university's test, tells The Scientist.
In another screening application, University of Illinois chemist Martin Burke laid out plans to use a new test designed by himself and colleagues on more than 50,000 students as they return to campus in the fall. "We imagine this will be just part of their orientation," Burke said during a webinar on June 16. "So you get your housing information, your dining card, your ID card, and you also submit your saliva sample."
Despite the ease of sampling, saliva is not without its challenges. For PCR, the virus's RNA must first be reverse transcribed into DNA, and saliva contains enzymes that chew up nucleic acids and inhibitors that interfere with the DNA amplification process used to detect the virus. As a result, saliva must often be purified before the DNA can be amplified. "It's kind of a dogma . . . that you have to start with absolutely pure DNA or RNA," Meagher says.
LAMP in the spotlight
While quantitative reverse transcription PCR remains the most-used method to diagnose Covid-19 regardless of how the sample is collected, many of the bottlenecks in the pipeline stem from the high cost and low scalability of the approach, says Meagher. A single benchtop thermal cycler, the machine that controls the temperature changes during the PCR, typically costs around $25,000 and can usually only run between 96 and 384 samples at a time.
To address these challenges, a team of researchers at Columbia University recently modified a LAMP protocol used by their fertility clinic to identify abnormalities in chromosome numbers in human embryos. Rather than count chromosomes, the tool now detects the presence of SARS-CoV-2 RNA in saliva in as little as half an hour, changing the color of the sample from red to yellow when the virus is present.
"For the Covid-19 pandemic, it was really felt that the big challenge is to try to get the test out to where it's needed," says Zev Williams, a reproductive endocrinologist who helped develop the test. "Perfectly accurate tests that take days to come back with results and can only be given to a small number of patients are not sufficient to address the problem."
The team's new test is one of several using LAMP. Unlike PCR, which creates new copies of DNA through cyclical temperature changes, LAMP reactions take place at a consistent 63°C, eliminating the need for complex machinery. The chemicals used in the reaction are also more robust against the enzymes and inhibitors in saliva, doing away with the need to purify each sample.
 "Very early on, [LAMP] was recognized as something that would have applications for pathogen detection," says Meagher, who has himself used LAMP to study Zika and Ebola. "You have the potential for a greatly simplified 'sample in-answer out' kind of assay, which is much harder to achieve with PCR."
LAMP has traditionally been deployed in resource poor countries because it requires less power and equipment than PCR. But under the current pandemic, the whole world has become somewhat resource limited, and Meagher stresses that LAMP can be just as useful in diagnosing emerging diseases in first-world nations such as the US. During Florida's Zika outbreak in 2016, diagnostic tests could sometimes take as long as eight weeks to process, Meagher says, and "a technique like LAMP would have made a huge difference in the United States as well."
See "Quick and Cheap Zika Detection"
A report detailing Williams's modified LAMP assay, published June 16 on the preprint server medRxiv, describe a "high-performance LAMP" with 1,000 times the sensitivity of a standard LAMP assay, capable of detecting a single copy of the SARS-CoV-2 virus in as little as one microliter of saliva. When compared to other methods such as RT-quantitative PCR, Williams says, the new LAMP assay can sometimes be even more sensitive.
Williams envisions his test being used to conduct mass screenings at airports, offices, and schools to identify even asymptomatic patients with a low viral load. "It's not the person who's sick and in the hospital" who needs treatment, he stresses. "We need to treat the person who's well and riding the subway and transmitting it." The team is currently applying for emergency use authorization.
Alan Wells, the medical director of the University of Pittsburgh's UPMC Clinical Laboratories, expressed skepticism that any LAMP assay could be as sensitive as traditional PCR. "While they could be as sensitive in theory, in practice I have yet to see it," Wells tells The Scientist, adding that LAMP assays are inherently less efficient at amplifying DNA than PCR is. "If this test becomes available . . . many of us will acquire and test it. If it works out as they say, that could be a wonderful addition."
While Williams's paper has not yet undergone peer review, it is far from the only research suggesting that LAMP may be as sensitive as PCR-based methods. Several projects have already begun processing samples using LAMP. The pilot study in England, for example, is using its own LAMP protocol, and researchers at Beaumont Health, Michigan's largest healthcare system, have also announced a new LAMP test for processing saliva samples.  
"I have never felt so much stress or pressure in my life," Brooks says, adding that he has frequently worked 22-hour days in the last four months to bring his spit test to market. "We've all taken it to another level because of its importance. I hope we never see anything like it in our lifetime again."

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