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I think to authors are overly optimistic in our ability detecting highly dilute samples. Pooling studies in Dengue Fever using RT-PCR found sensitivity fell fas

I think to authors are overly optimistic in our ability detecting highly dilute samples. Pooling studies in Dengue Fever using RT-PCR found sensitivity fell faster than could be explained by dilution effects alone. At 100:1 or 20:1 pooling the false negative rate will be unacceptable. Edit: false positive -> false negative

virusduck 6y ago
False negative rate maybe? In general, titers of the virus in NP swabs should be high enough to withstand 10, maybe 20-plex pooling. However, as you allude to, there are plenty of other effects that can influence PCR. If there is one sample, for example, that contains a PCR inhibitor, like heme, it may then inhibit all the samples it is pooled with. Additionally, swabs are not just swabs of virus, they also pull of highly variant amounts of bacteria and human material. These things may also affect the efficiency of PCR.
- bhickey 6y ago
  > False negative rate maybe? Yes, thanks, fixed. Check out Table 1 in this paper. It isn't very encouraging. https://onlinelibrary.wiley.com/doi/full/10.1002/jmv.25971 https://onlinelibrary.wiley.com/doi/full/10.1002/jmv.25971
cjlars 6y ago
This is very, very, very easy to test experimentally though, right?
- xster 6y ago
  It's already been done no? That's how Wuhan tested 11 million residents in May.
  - pmiller2 6y ago
    I'm not saying you're wrong, but, if that's the case, why haven't we used the same technique already to ramp up testing?
    - bhickey 6y ago
      This has been used at the University of Nebraska Medical Center for a few months now. They do 5-way pooling.
    - FuckButtons 6y ago
      The interview covers this to an extent, the reason is a lack of appreciation of the potential and an overestimation of the complexity. It’s a somewhat complicated thing to multiplex all the samples correctly, it’s too complex a scheme for an individual lab scientist to do, it either requires a robot or some form of semi automation, I think in Rwanda they were using a smartphone app to tell the scientists which batch to put each sample in. I think that if you could convince the worlds public health bodies, and we can get saliva sampling off the ground in combination you could get to the levels of testing needed to actually do national level surveillance.
  - bhickey 6y ago
    Wuhan tested in batches of 10.
    - jmpman 6y ago
      Do they stick the same q-tip up everyone’s nose?
      - bhickey 6y ago
        That would be a very effective way to achieve herd immunity quickly. Typically the swab is stored in a liquid viral transport medium. You perform a RNA extraction on this liquid, pool the product and run RT-PCR on it.

virusduck 6y ago

False negative rate maybe? In general, titers of the virus in NP swabs should be high enough to withstand 10, maybe 20-plex pooling. However, as you allude to, there are plenty of other effects that can influence PCR. If there is one sample, for example, that contains a PCR inhibitor, like heme, it may then inhibit all the samples it is pooled with. Additionally, swabs are not just swabs of virus, they also pull of highly variant amounts of bacteria and human material. These things may also affect the efficiency of PCR.

bhickey 6y ago
> False negative rate maybe? Yes, thanks, fixed. Check out Table 1 in this paper. It isn't very encouraging. https://onlinelibrary.wiley.com/doi/full/10.1002/jmv.25971 https://onlinelibrary.wiley.com/doi/full/10.1002/jmv.25971

cjlars 6y ago

This is very, very, very easy to test experimentally though, right?

xster 6y ago
It's already been done no? That's how Wuhan tested 11 million residents in May.
- pmiller2 6y ago
  I'm not saying you're wrong, but, if that's the case, why haven't we used the same technique already to ramp up testing?
  - bhickey 6y ago
    This has been used at the University of Nebraska Medical Center for a few months now. They do 5-way pooling.
  - FuckButtons 6y ago
    The interview covers this to an extent, the reason is a lack of appreciation of the potential and an overestimation of the complexity. It’s a somewhat complicated thing to multiplex all the samples correctly, it’s too complex a scheme for an individual lab scientist to do, it either requires a robot or some form of semi automation, I think in Rwanda they were using a smartphone app to tell the scientists which batch to put each sample in. I think that if you could convince the worlds public health bodies, and we can get saliva sampling off the ground in combination you could get to the levels of testing needed to actually do national level surveillance.
- bhickey 6y ago
  Wuhan tested in batches of 10.
  - jmpman 6y ago
    Do they stick the same q-tip up everyone’s nose?
    - bhickey 6y ago
      That would be a very effective way to achieve herd immunity quickly. Typically the swab is stored in a liquid viral transport medium. You perform a RNA extraction on this liquid, pool the product and run RT-PCR on it.