10 August Progress in diagnostics LAMP and subgenomic RNA

1. RT-LAMP as a more rapid and cheap technique technique aq compared to RT-PCR: 2 papers

RT-LAMP combines reverse transcriptase, DNA polymerase, pH indicator, and six primers to amplify RNA templates causing a drop in pH and, thus, a color change from pink to yellow.

Since RT-LAMP involves both reverse transcription and DNA amplification at a constant temperature without the need for a PCR thermal cycler], it only requires simple equipment, namely benchtop centrifuges, heat blocks, and micropipettes. 

Both papers show that LAMP is very sensitive and specific.   Much, much better than the insensitive “rapid antigen tests”.  (see third paper)

I add some illustrations (from Wikipedia), showing the principles and a Table comparing different diagnostic methods.   

My questions to the experts in this audience :

• Is LAMP may the light at the end of the tunnel of the problems and bottlenecks we are still experiencing with the classical RT-PCR  
• Could LAMP easily be transferred to LMIC to make COVID diagnoses more affordable and accessible?  

2. Some information on Xpert for SARS-CoV-2:  also easy and reliable, but expensive as well.  See next 2 attachments.  

3.  Subgenomic RNA as a marker of “life” virus: a very recent paper in Emerging Infectious Diseases https://wwwnc.cdc.gov/eid/article/26/11/20-3219_article   shows a moderate correlation with viral culture.   Clearly, sgRNA may be a more sensitive marker. “Mild” patients with prolonged shedding of Nucleoprotein RNA do not show positivity in culture or sgRNA.

The sgRNA provides evidence of replicative intermediates of the virus, rather than residual viral RNA. 

Of 33 specimens tested for sgRNA and by virus culture, both tests showed positive results for 12 (36.4%) specimens, both tests showed negative results for 12 (36.4%), sgRNA showed positive results and culture was negative for 7 (21.2%) specimens, and culture was positive and sgRNA showed negative results for 2 (6.1%) samples.

Virus sgRNA was detectable in 18 (81.8%) of 22 specimens collected <8 days after symptom onset and in 1 (9.1%) of 11 specimens collected >9 days after onset of disease (p = 0.0003 by χ2 test with Yates correction) (Figure 2).  

Figure 2. Severe acute respiratory syndrome coronavirus 2 viral RNA load, virus culture, and subgenomic virus RNA (sgRNA) in relation to days after onset of illness for patients with mild coronavirus disease, Hong Kong. Red indicates culture and sgRNA positive, green indicates culture and sgRNA negative, yellow indicates culture positive and sgRNA negative, brown indicates culture negative and sgRNA positive, and gray indicates culture positive and no sgRNA data (because of insufficient specimen).