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[SANET-MG] field testing GM bacteria in Wisconsin
August 12, 2005
Prof. Joe Cummins
“Field testing Erwinia carotovora bacteria containing antibiotic
resistance genes”
The University of Wisconsin is preparing to release genetically modified
(GM) Erwinia carotovora bacteria containing antibiotic resistance marker
genes in environmental field tests. The United States department of
Agriculture (USDA) Animal and Plant Health Inspection Service (APHIS)
has prepared a docket with background information on the release and
there has been a public comment period ending September 15,2005.. The
tests are to be conducted at the University of Wisconsin Experiment
Station near Hancock, Wisconsin. The docket is located as described in
reference (1). The release to the environment of a bacterial strains
each containing a different antibiotic should be a matter of grave
concern and a strong public response is in order.
The purpose of the field trial seems to be simply experimental with no
potential commercial application of the genetically modified bacteria.
Erwinia carotovora causes a rot disease in potatoes. The experiment is
designed to test the fitness of bacteria that have had genes known to be
associated with pathogenesis which have been disrupted by the insertion
in those genes of the antibiotic resistance genes. The antibiotic
resistance genes not only eliminate the function of the pathogenesis
gene but they also serve as selectable markers to identify the modified
bacteria. The antibiotic resistance genes are precisely inserted into
the target pathogenesis gene by adding short DNA tails to the resistance
genes that are homologous to the target pathogenesis gene. The
pathogenesis gene target is Hrp which controls secretion of virulence
proteins into the host plant. The green fluorescence protein was
inserted as a visual marker to identify potatoes bearing the modified
bacteria (2).
The field test involves four GM mutant strains (WPP60,WPP198,WPP195 and
WPP40. Strains WP60 was created by inserting the
streptomycin/spectomycin genes into the hrcC gene while WPP98 had
chloramphenicol resistance gene inserted into hrpL gene.WPP195 was
created by deleting the hrpN gene and inserting the choramphenicol
resistance gene in its place. Strain WPP was created by inserting the
kanamycin resistance gene into the OutD gene that is required for a
functioning protein secretion system in the bacterium.; The mutant
cannot secrete plant cell wall degrading enzymes. Inserting antibiotic
resistance genes into the hrp genes interferes with pathogenisis.
However only strain WPP40 that eliminates the OutD gene is reduced in
virulence in a direct assay while the other three strains are not
reduced in virulence, but in spite of that they are carried in the field
experiment (2).
The field tests were not expected to impact agricultural practice
because “the test is solely for research purposes” even though Erwinia
carotovora effects potato, cucumber, capsicum, turnip, brussel sprouts ,
carrots and celery, all of which are grown in the general area of the
test site. .Because the antibiotic resistance genes are not being
selected by antibiotic application they are not believed to be a concern
according to the applicants. The proposal believes that the antibiotic
resistance genes cannot be transferred to bacteria effecting humans (as
pointed out below that belief is not reasonable). The applicants justify
that position by judgments from the US Food and Drug Administration that
antibiotic resistance is already prevalent in bacteria effecting humans
and in soil born bacteria. Even though the soil is sandy and porous in
the area of the test site spread of the GM bacteria is not expected nor
tested for(2). In general the application focuses on the benefits of the
experiments and ignores any evidence that might delay the experiment..
The most obvious question is why release so many strains that are
genetically modified but still virulent? That procedure provides high
risk with little or no benefit.
The belief expressed in the proposal, that GM Erwinia carotovora would
not transfer antibiotic resistance genes to other soil bacteria seems
inconsistent with the scientific literature. Horizontal gene transfer
was commonplace in Erwinia and Klebsiella and further that transfer
could extend to Salmonella, Shigella and E. coli (3).Presumabley the GM
bacteria on surfaces of food crops could transfer antibiotic resistance
gene to enteric bacteria during ingestion by humans or animals. Bacteria
phage were found to transfer genes between Erwinia species
(4).Transposons capable of mobilizing antibiotic resistance genes have
been identified in Erwinia carotovora (5).There is clear evidence that
Erwinia is capapble of moving resistance genes among soil Erwinia
species, between other soil bacterial species and among enteric bacteria.
A number of GM microbes bearing antibiotic resistance markers have been
released commercially in the United States , these include
Sinorhizobium, Agrobacterium and Psueomonas (6). Little effort seems to
have been put forth to monitor the consequences of these releases. The
hazards of horizontal gene transfer have been discussed in depth by Dr.
MaeWan Ho (7,8).
The main problem with the proposed field test of GM Erwinia carotovara
is the failure to monitor the spread of the modified bacteria and the
horizontal spread of the genes with which they have been modified. The
release of virulent GM Erwinia does not appear to provide any benefit in
the face of the risk of spreading the antibiotic genes.
Reference
1. University of Wisconsin-Madison Availability of Environmental
Assessment for Field Tests of Genetically Engineered Erwinia carotovora
Agency Document Number APHIS-2005-0069-0001 Comment Period End Date
(mm/dd/yyyy): 09-12-2005
http://docket.epa.gov/edkfed/do/EDKStaffCollectionDetailView?objectId=0b0007d48094780b
2. USDA/APHIS Environmental Assessment in response to permit application
(05-097-01r) received from University of Wisconsin for field testing of
genetically engineered strains of bacterium, Erwinia carotovora subsp.
Carotovora 2005
http://docket.epa.gov/edkfed/do/EDKStaffAttachDownloadPDF?objectId=090007d48094780e
3 Mulec J, Starcic M and Zgur-Bertok D. F-like plasmid sequences in
enteric bacteria of diverse origin, with implication of horizontal
transfer and plasmid host range. Curr Microbiol. 2002 Apr;44(4):231-5
4. Chatterjee AK, Ross LM, McEvoy JL and Thurn KK.
pULB113, an RP4::mini-Mu plasmid, mediates chromosomal mobilization and
R-prime formation in Erwinia amylovora, Erwinia chrysanthemi, and
subspecies of Erwinia carotovora. Appl Environ Microbiol. 1985 Jul;50(1):1-9
5. Kotoujansky A, Lemattre M and Boistard P. Utilization of a
thermosensitive episome bearing transposon TN10 to isolate Hfr donor
strains of Erwinia carotovora subsp. Chrysanthemi J Bacteriol. 1982
Apr;150(1):122-31
6. Cummins,J. GM Microbes invade North America Sience in Society 2003, 19.39
7. Ho,MW Horizontal gene transfer happens-II ISIS Report 2001
http://www.i-sis.org.uk/
8.Ho,MW Recent evidence confirms risks of horizontal gene transfer ISIS
contribution to ACNFP/Food Standards Agency open meeting 2002
http://www.i-sis.org.uk/
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