BEE-L members:

I received the following from Dr. Diana L. Cox-Foster (who is not a 
subscriber to BEE-L, hence she cannot post to the list) with a request to 
forward it to BEE-L.

Sincerely,
Aaron Morris

An open letter regarding Colony Collapse Disorder and science
September 25, 2007

We encourage constructive discussion and welcome re-examination of our 
findings
and conclusions regarding CCD, as reported in Science magazine. However, 
common
sense and ethical rules in science require that such critiques are made with
full disclosure of methods and conflict of interest. The beekeeping 
community
should accept no less, and we feel that this has sometimes been overlooked 
in
the case of CCD discussions. In particular, Dr. Jerry Bromenshenk of Bee 
Alert
Technology, Inc. has commented extensively on specific strategies for 
assessing
the causes of Colony Collapse Disorder and on our recently published study 
on
CCD correlates. He has been interviewed frequently in the media and has
provided quotes to Science magazine as well as other media that have been
widely cited. Among those comments were vague statements to the effect that 
he
has evidence relevant to the distribution of Israeli Acute Paralysis Virus
(IAPV) in CCD and healthy bee colonies. These claims are fleshed out in his
open letter of Sept. 10 (published online at www.beeculture.com), but are
missing critical elements (i.e., data) for evaluation.

The work reported in Science magazine was performed by academic, federal and
state investigators who have no vested interest in its outcome. None of us 
have
patents or intellectual property of any kind that would benefit from finding
any result. We have no instrument to sell or rent. We do not offer
fee-for-service screening. We have not applied for patents, grants or
contracts. The USDA has an internal budget, and Penn State has modest 
support
from USDA and the Pennsylvania Department of Agriculture. The Columbia site
volunteered hundreds of thousands of dollars in supplies and personnel time
because, as faculty in a School of Public Health, they were persuaded by 
Drs.
Cox-Foster and Pettis that the need was urgent and that this was an 
appropriate
area for them to contribute to global health and welfare.

Our survey was initially performed using pyrosequencing. Although this
technology is not yet widely available it is based on standard molecular
biological techniques that are accepted by the scientific community. Recent
prominent examples that include sequencing of the genome of a Neanderthal 
and
of James Watson, Nobel laureate and co-discover of the DNA double helix. We
followed our initial work with well established testing methods that 
included
culture, manual spore counts, polymerase chain reaction (PCR) measurements 
of
multiple gene regions of the disease organisms, and extensive sequencing by
conventional means. The PCR measurements were performed in two independent
laboratories by scientists blind to sample codes. Thus, we have reported
results that are consistent and objective.  Contrary to some claims on the 
web,
these analyses were performed on hundreds of bee samples from multiple
operations having CCD and healthy bees from different origins.

PCR has become one of the best established tools in microbiology. Since Kary
Mullis received the Nobel Prize for discovering PCR in 1993, it has 
literally
revolutionized management of outbreaks of infectious disease and 
environmental
surveillance. The method is sensitive, specific, and inexpensive. Hundreds 
of
assays have been approved by the FDA and other regulatory bodies concerned 
with
monitoring disease, blood products, and food and water safety. The term
"PCR" retrieves more than 299,000 articles in PubMed, the database of the
National Library of Medicine. In contrast, IVDS is a method available only
through Dr. Bromenshenk and colleagues. The term IVDS retrieves no relevant
articles in PubMed (only references to "in vitro devices" and
"intravertebral disks"). A search for IVDS on the more inclusive
'Scopus' database yields 5 relevant publications authored by IVDS developer
Charles Wick since 1999 (four involving sorting of a single bacteriophage
isolate, and one involving bacteria) and 7 patent citations. A query using
"PCR" in the Scopus database retrieves 265,394 research articles and
252,155 patent citations.

PCR is a widely used technique in bee pathology and disease, that relies on
specific matching between diagnostic 'primers' and any DNA (or processed
RNA) source. This technique has worldwide acceptance as a regulatory and
research tool. Given the sequencing of the bee genome as well as the genomes 
of
the major bacterial, fungal, and viral pathogens of bees, the potential 
targets
for PCR-based diagnostics are almost limitless, allowing multiple tests to
confirm or refute the presence of pathogens. Most importantly, PCR is easily
followed by the determination of exact DNA/RNA sequences of the targeted
pathogen or parasite. There is simply nothing more diagnostic of an organism
than its genetic code.

For several months, we have encouraged the primaries behind IVDS (Dr.
Bromenshenk at Bee Alert Technologies, Dr. Charles Wick at the U.S. Army's
Edgewood Chemical and Biological Center, and Mr. David Wick, the President 
and
CEO of Biological Virus Screening, inc., the business arm of IVDS,
http://www.bvs-inc.us/) to carry out experiments showing that IVDS is 
specific,
sensitive and reliable with respect to bee viruses. Given the identification 
of
over 19 different viruses in honey bees worldwide, it is not an easy matter 
to
confirm specificity. These encouragements have been made in good faith, 
since
it is always better to have alternate technologies on the table. Based on
published research, IVDS specificity has not been adequately tested against 
bee
viruses. Such tests could start by analyzing pure, known viruses and should 
be
followed by attempts to recover viral signals from bees with known 
infections.
Such evaluations have been completed on every diagnostic test for bee 
viruses
since the 1970's, and they are also required for IVDS before adopting this
technology.  We and other researchers are concerned that the tight size
variation of most bee viruses (28-30 nm) and the relatively large error
implicit in IVDS (+/- 4 nm) precludes a completely specific diagnostic. The
next stage is to assess sensitivity. PCR is currently considered to be the 
most
sensitive method available for diagnostics, capable of capturing even single
nucleic acid molecules.  Such sensitivity may not be needed here; however, 
we
must at least know the extent to which IVDS is able to distinguish viral
particles from one another and from the natural material found in ground up
samples. From the available postings (again restricted to
http://www.bvs-inc.us/), it is not apparent that the required calibration 
tests
are planned, let alone conducted). These are urgently needed prior to using
IVDS to weigh in on viral distributions in bees in different regions of the 
US
or the world. It is conceivable that IVDS may prove useful; and we encourage
the development of new and complementary diagnostic techniques. However, it 
is
premature to state that IVDS has ruled out a pathogen found by accepted 
methods.

Dr. Bromenshenk and colleagues claim to have found 4-6 new viruses in
Australian and American based on IVDS size peaks. This is intriguing, and we
and others look forward to seeing the data when it is reported for peer 
review.
 According to their own press reports, we understand that they found Nosema
ceranae and an unidentified iflavirus in CCD bees
(http://pub.ucsf.edu/today/cache/feature/200704251.html). IAPV and KBV are
dicistroviruses, not iflaviruses. Dicistroviruses are easily distinguished 
from
iflaviruses using molecular methods. Discistroviruses have an intergenic 
region
and  encode structural proteins at the 3'-end rather than the 5' end of
their genomes.

It has been stated in this and other forums that we have been secretive. Our
initial findings were presented in meetings of the Institute of Medicine in
Washington DC in April 2007, and at the annual meeting of the Regional 
Centers
of Excellence in Biodefense and Emerging Infectious Diseases in St Louis In
April 2007. IAPV was identified as a candidate in these presentations.
Regretfully, we could not do more than list all candidates prior to 
conducting
a more detailed analysis and submitting the data for rigorous peer review at
Science magazine. These experiments added months to the process but served 
to
downgrade the importance of tens of candidate pathogens and parasites. We 
hope
our paper, the first published on CCD research, helps pave the way for
additional field, lab and survey approaches to understanding CCD. Many
researchers have become attracted to this problem, a good thing for 
beekeepers
and for the resolution of this and other bee maladies.

We have not claimed that IAPV is the cause of CCD. We have said that IAPV is
highly associated with CCD and should be pursued as a potential trigger for
disease. We also have not said that bee imports from Australia should be
banned; in fact, this is not a decision to be made by any of the members of
this research group. Indeed, the fact that IAPV has now been found in the 
US,
Australia, Israel and China (royal jelly samples) means that the virus may
already be globally distributed. The IAPV finding does, however, provide the
community with an opportunity to test for the presence of this virus in
colonies, imports and exports, using inexpensive, specific assays. No one 
would
argue that unwitting transfer of infected materials-whether bees, food 
products
or animals-is a good thing. Thus, we have published detailed descriptions of
the PCR sets we used and have already provided standards at no cost to
laboratories who wish to set up their own tests. This is the same policy the
Columbia group (then at the University of California) applied when West Nile
virus was discovered in 1999, and in subsequent work that has received less
media attention. The bee industry deserves state-of-the-art accurate science
and full disclosure.

W. Ian Lipkin
Thomas Briese
Sean Conlan
Gustavo Palacios
Columbia University

Diana Cox-Foster
Edward Holmes
Pennsylvania State University

Dennis van Engelsdorp
Pennsylvania Department of Agriculture

Jay Evans
Jeffrey Pettis
United States Department of Agriculture

Nancy Moran
University of Arizona, Tucson 

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