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From:
Jerry Bromenshenk <[log in to unmask]>
Reply To:
Informed Discussion of Beekeeping Issues and Bee Biology <[log in to unmask]>
Date:
Fri, 22 Mar 2013 21:25:28 -0400
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Juanese asked about viruses, nosema in failing bee colonies.
 
Yes, its all available in our PLOS ONE paper, Iridovirus and Microsporidian 
 Linked to Honey Bee Colony Decline, October, 2010.
 
As you know, we have had some very vocal critics, but we're  now publishing 
a rebuttal that says in essence - we did it right, the  others have it 
wrong.  This rebuttal has taken some time due to false  claims that we had to 
deal with over past two years.  Our data has now been  carefully examined by 
two independent groups and found to be solid and complete  and appropriately 
analyzed.  That took some time.  
 
There is a paper by another research group who looked at 3  of our samples 
and state that they did not find nosema nor evidence of a DNA,  
iridescent-like virus.  Problem is, they got our samples without our  knowledge..    
Army sent raw data for three samples to one group,  who without asking 
permission, passed them to another group, who then  re-analyzed and published a 
quick paper, making over-stated claims that their  analysis differed, and as 
such, we (Army) was shown to be wrong, and that their  paper should serve as a 
warning.
 
Here's our summary, which is a very small part of our  full  rebuttal:
 
Army  sent out three samples as per request of another group, who  passed 
them on.  These samples were carefully picked and sent as blind  samples - 
the recipients had no knowledge of the provenance. 
 
One sample by Army ABOID analysis had no Nosema and no IIV-like virus  (it 
was a true control)
One sample had very small amounts of Nosema and IIV-like virus (it served  
to test analysis  detection limits,
Only one sample had both pathogens (it was the positive control - they  
should have found both pathogens)
 
They  re-analyzed these 3, got their paper published, without asking  about 
the nature of the three samples that they re-examined.  
 
They then wrote a paper saying - no nosema, no IIV virus.  They then  
alleged that they were right, Army was wrong.
 
Interestingly, they did not dispute all of the small, RNA viruses that we  
found -they also found them.  
 
Again, they claimed none of the samples had Nosema or an IIV-like  virus.
 
They claimed in their paper that we failed to look at bee proteins, which  
is patently untrue - we looked at all bee proteins and a wide array of honey 
 bee, insect, and plant pathogens, which was clearly stated in our article. 
  We chose not to include the bee body proteins in our statistical 
analysis,  choosing instead to focus on the pathogens.  Its not that we didn't look 
at  them - they were inappropriate for the analysis.
 
There was no reason to include the proteins of the bee host in a  
statistical analysis of pathogen proteins in CCD colonies.  There may be  another 
paper there, if we chose to see if somehow the host's proteins were  altered by 
the pathogens, but that wasn't out purpose.
 
Where they went wrong is:
 
1) analyzing samples and writing a paper without asking about the nature of 
 the three samples - which they said represented all of our samples, which 
is  incorrect - they got carefully picked samples to test whether others 
could  properly classify them (it was an intentional test by Army and they took 
the  bait)
2) stating that they neither found the IIV-like virus or Nosema   Here's 
where they went over the cliff.  
 
We confirmed the Nosema by both PCR and microscopy, in the two of the three 
 samples that had Nosema,  
 
To be clear, we knew other groups did not agree with our finds.  Army  
chose to hand-pick three samples to sent to anyone wanting to re-analyze.   
These  were NOT picked to represent out full data set, they were sent to  
determine whether the other groups could properly categorize the  proteins  in the 
three samples.  One blank or control, one very low level infection  that 
would test limits of detection, and one sample with both pathogens, the  only 
sample that Army expected anyone else to find Nosema and virus in.
 
So their abstract proclaims that the three samples had no Nosema, no  
IIV-like virus, and that these three samples were representative of our CCD  
samples - wrong on all accounts - too bad they can't read.  
 
There is no doubt that two of the three samples had Nosema.  Army  
proteomics, MSU PRC, and MSU and UM microscopy confirmed.  In other words,  two labs 
confirmed the Nosema using non-proteomics analysis. As you all know,  one 
can see Nosema spores under a microscope, and adding PCR brings a genomic  
analysis to bear on the question.  Nosema in two of the three samples were  
confirmed by three independent methods and two different institutions.   There 
is no doubt that those samples had Nosema, and if they had Nosema, then a  
proteomics analysis should have found proteins from Nosema.  Army's  
proteomics did, theirs didn't.
 
 
  If  they couldn't find Nosema proteins, then they have no  justification 
for claiming that Army analysis was wrong.  Their approach  either lacked 
the sensitivity or specificity to detect, not Army's.  If  they can't find 
Nosema, then any claims about whether IIV was present is  mute.  It like being 
so blind as to not see the elephant in the room, but  arguing that since 
they couldn't see the elephant, there certainly couldn't have  been a mouse in 
the opposite corner.  If you can't see the elephant, you  can't see the 
mouse.  If others can see the elephant, you have to accept  there may have been 
a mouse in the room.
 
3) assuming that analysis algorithm they used was comparable to the custom  
ABOID software developed
by the Army, and licensed last year to Sage-N-Research for intensively  
computational protein analysis.  Sage-N-Research and Army conducted many  tests 
of the accuracy of the Army software over past few years.  They were  
engaged in this testing when our paper emerged.  Sage-N licensed Army ABOID  
software based on its unique capabilities and proven ability to detect microbes  
and Id to strain, not just species.
 
So, our formal rebuttal  verifies our original results and points out  why 
our detractors failed to find the same results.  
 
That said, we have seen a lot of viruses in collapsing bees plus  nosema 
ceranae, some nosema apis, but varroa only in a subset of the samples we  
analyzed over several years.  
 
Jerry
 

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