Eighth International Molecular Plant-Microbe Interactions Congress
Eighth International IS-MPMI Congress Program
IS-MPMI Members Choose Your Logo!
IS-MPMI Publishes 1996 Proceedings
Calendar of Events
Note From the Editors
As you know, the 8th International Congress (and concurrent 7th Annual Gatlinburg Symposium) will be held July 14-19, in Knoxville, Tennessee. The meeting will emphasize the impact of molecular biology on all aspects of plant-microbe interactions.
THERE IS STILL TIME TO SUBMIT YOUR ABSTRACTS! Abstracts will be accepted through April although the original deadline was March 15th. However, we cannot ensure the publication of your abstract unless it is received by the end of April.
If you have not yet registered, we urge you to send in your materials as soon as possible. You can receive registration materials by contacting: UT Conferences, The University of Tennessee, P.O. Box 2648, Knoxville, TN 37901 USA, phone: 615/974-0250, fax: 615/974-0264, http://www.ce.utk.edu/departments/conference/confhome.html E-mail: firstname.lastname@example.org. For your reference, the IS-MPMI Congress program has been printed in this newsletter and begins on page six.
When you receive your registration materials there are a few points of clarification. First, when submitting fees, your check should be made payable to: "The University of Tennessee". Second, a block of rooms at the Holiday Inn has been reserved for the meeting. The rooms have been reserved under the name "Molecular Plant-Microbe Interactions Meeting". Make sure you give this title for the meeting or they may incorrectly tell you that the rooms have all been booked. Third, the tours mentioned in the registration materials are quickly becoming booked so, again, make sure you register as soon as possible.
Finally, the University of Tennessee has agreed to
offer 2 graduate hours of credit for attendance at the Symposium.
For more information contact, Dr. Beth Mullin, Department of Botany,
University of Tennessee, Knoxville, TN, 37996-1100, phone: 423/974-6203,
fax: 423/974-0978, E-mail: email@example.com.
See you in Tennessee!
IS-MPMI Is Growing
This past year has been a busy and productive one for our Society. It's very exciting to see our membership growing and our member services expanding. Over the past year, several changes have been made to better accommodate the changing needs of our members and I would like to take this opportunity to review these changes with you.
The Senior Editorial board of MPMI has taken several steps to improve the quality of our Society's journal. MPMI will now be published nine times a year instead of six. The price of publishing color plates has been lowered and the editorial content is being expanded to include reviews, discussions, etc. An increased emphasis has been placed on the plant side of the interaction and a new senior editor has been added. Taken together, these changes will help attract additional quality papers to MPMI, improve the turnaround time for printing articles, and attract a broader reading audience.
Recruiting New Members
Members Alan Collmer and Barry Rolfe have produced our first membership brochure. The brochure and a cover letter which discusses the benefits of IS-MPMI membership were sent to approximately 2,000 prospects. We will also send membership promotional materials to all non-member attendees of the Knoxville meeting. Our membership is at an all-time high with 478 members.
This past year we also developed an Internet homepage for our Society at http://www.scisoc.org/ismpmi. It currently includes the listing of recently accepted abstracts for MPMI, past issues of the IS-MPMI Reporter, updates on the Knoxville meeting and links to other information on the Internet of interest to members.
Thanks to the work of Sally Leong and Ulla Bonas we produced our first two issues of the IS-MPMI Reporter newsletter in 1995 and we are currently on-schedule to produce three issues in 1996.
Updating The Society's By-laws
Our growing and changing Society has prompted a review of our by-laws. The board is currently examining and revising the by-laws.
A copy of the revised by-laws for the society will be mailed to all members in May. The by-laws, as amended, will be subject to approval by a vote at the business meeting at the IS-MPMI meeting in Knoxville in July.
Increased Headquarter's Resources
In addition to our board and volunteer members, the headquarter's staff has assisted us in accomplishing the goals and projects we had set for 1995 and 1996.
In January, some members of the APS and IS-MPMI boards met to discuss issues of mutual interest to both societies including a mechanism for communication especially as it relates to changes to the MPMI journal. Although there was no formal introduction and acceptance of future collaboration of projects, it was recognized that both societies could benefit by shared staff resources and overall association management.
This summer the headquarter's editorial and production staff will be used in the publishing of the proceedings of the 1996 International Molecular Plant-Microbe Interactions Congress under the IS-MPMI publishing imprint.
IS-MPMI 1998 Meeting Site
The Board of Directors will review potential sites outside of North America for the 1998 IS-MPMI meeting at their board meeting in Knoxville on Sunday, July 14, 1996. Individuals who are interested in hosting the 1998 meeting are invited to submit a letter of interest to me c/o Department of Microbiology, Box 357242, University of Washington, Seattle, WA 98195-7242, USA. Phone: (206) 543-0255; fax: (206) 543-8297; E-mail: firstname.lastname@example.org.
To simplify the planning and reduce the amount of work of the host organizing committee, the headquarters staff is prepared to work with the IS-MPMI Committee that agrees to host the 1998 meeting. In the past, we have depended on member volunteers to oversee all the details of executing a meeting. If requested, the headquarters professional meeting planners are prepared to assume the responsibilities for organizing the meeting, and IS-MPMI members will be responsible only for the development of the programming aspects of the meeting.
It has been an exciting year and there is still much more to be done. We want to hear from you. After all this is your Society and we want you to help shape its future. Please contact me or any of our Board members with your ideas, thoughts, etc. In the meantime, I look forward to seeing you in Tennessee.
The REMI technique (restriction enzyme-mediated integration) was introduced in Saccharomyces cerevisiae (1). It was quickly employed in Dictyostelium discoideum (2) and has been extensively used in this organism for producing tagged mutations. Similar efforts, published (3-7) and unpublished (8-15), have started for a number of filamentous fungi and that work is the subject of this review.
How REMI works
The REMI technique as introduced in yeast (Fig. 1A) differs from a normal transformation in that the transforming DNA is linearized with a restriction enzyme and an aliquot of the same restriction enzyme is added to the transformation mix. These components make their way to the nucleus where the restriction enzyme cuts genomic DNA at its recognition site. This break in the genomic DNA serves as the site of integration for the transforming vector. The compatible cohesive ends between the vector and the restricted genomic DNA catalyze the integration event, i. e. a Bam-cut vector ligates in vivo to the Bam-cut genomic DNA with two Bam sites being generated.
Some integration events in fungi do not retain the vector ends and/or the genomic DNA ends in their original form and thus restriction site restoration fails to occur (Fig1b). In Magnaporthe grisea we have clear evidence that, despite the failure to regenerate this restriction site, integration does indeed occur at a genomic site corresponding to the enzyme added to the transformation. This difference in integration events need not be confusing. The fundamental function of restriction enzymes is the production of double strand breaks in the genomic DNA which then serve as the site of integration for transforming DNA. The nature of the recombination system in the organism and the fate of free DNA ends during transformation determine whether the compatible cohesive ends will be used and preserved in the integration event.
If double strand breaks are limiting in transformation then it follows that REMI should increase transformation efficiencies and it does. Transformation rates often increase 5- to 10-fold and increases as great as 100-fold have been reported. Enzymes are added immediately before treating cells with the agent(polyethylene glycol/calcium, lithium acetate, or electroporation) that causes DNA to enter the cells. Many different enzymes, including those with four- and six-basepair recognition sites, have this effect (usually 10-50 units/transformation). The optimum enzyme concentration, however, must be determined empirically and the optimum is often a sharp peak. Restriction enzymes can also increase transformation efficiency even if no correspondence exists between the enzyme used to cut the vector and the enzyme used in the transformation and even with uncut transforming DNA. We routinely add 10 units of Bcl I to transformations with uncut DNA to get a 5-fold increase in transformation efficiency. (Bcl I is inexpensive and also is sensitive to dam methylation and won't restrict DNA prepared from dam+ E. coli.) Restriction enzymes can also decrease the number of transformants, probably because genomic DNA cutting overwhelms cellular repair systems and leads to cell death. This result indicates that the enzyme finds DNA and functions in the nucleus and suggests that a lower concentration could actually increase efficiency. The most important benefit of REMI for some organisms, therefore, may be simply to make transformation better.
Tagging by transformation--Why REMI?
First, consider that the perfect tagging agent for filamentous fungi would integrate randomly, would involve only a single vector molecule at a single genomic site, would occur without any damage to the vector or the genomic DNA, would allow facile recovery of the vector and some flanking sequences and would cause no additional untagged mutations. Also consider that since integrative transformation is the rule in filamentous fungi, normal transformation could and has been used to tag mutations and facilitate subsequent gene cloning. Why then have researchers recently chosen REMI transformation versus normal transformation? In large part the decision to use REMI was based on a sense (rather than extensive experimental data) that REMI transformation would be closer to the perfect tagging agent than normal transformation and also a sense that the success of the D. discodeum tagging efforts could be emulated in fungi. Some points worth considering when employing REMI are:
Randomness. A rigorous answer about the randomness of integration requires enormous efforts so most workers plunge ahead without answering the question. When seeking a single gene the point is particularly critical, but work to date offers no clear answer for filamentous fungi. The Tox1 locus of C. heterostrophus locus was easily tagged (2/1310 transformants) (3), though this is a very large gene. On the other hand, we have failed to find a single tagged melanin biosynthetic gene in M. grisea among 5500 transformants even though the pathway contains at least four genes. Also, we have independently tagged three different genes twice among 13 genes identified. Nevertheless, workers searching for broad classes of mutants (e.g. pathogenicity or developmental genes) (6, 7, 10, 11, 12, 13, 14) have found a wide range of mutants so clearly hot spots are not interfering with the utility of the technique. Even if restriction enzymes cut randomly in the genome, a spectrum of enzymes must be used to insure a site exists in every gene of interest. Theoretically, enzymes with four basepair recognition sites would provide the best coverage.
Complexity of integration events. Researchers can rigorously examine the nature of integration events more easily than the randomness question and these investigations are probably worthwhile since simple integration events make for easier downstream cloning. Several workers (6, 13, 14) report that including restriction enzymes minimizes tandem integrations thereby maximizing single vector integration events. This phenomenon might be caused by restriction enzyme cutting of concatemers to maintain a pool of single vector molecules for integration. Simple integration events are especially useful for fungi without genetics. Recovery of a single integrated vector with flanking DNA from both sides of the integration results in a pre-made gene disruption vector (see Fig 1). Retransformation of a wildtype strain can then verify that the original insertion caused the detected mutation.
Modification of vector and genomic DNA. Transformation events in fungi have sometimes lead to deletions. Indeed, REMI might be useful in stimulating deletions (if in fact they were desired). Deletions at the insertion point can complicate cloning since the DNA proximal to the inserted vector might not contain the gene responsible for the mutant phenotype. We have found occasional deletions of genomic DNA as big as several kb at the insertion points. We have also observed in M. grisea that most linearized transforming vectors have a few to several hundred basepairs removed. This may occur to a lesser extent in other strains of M. grisea (10). Untagged mutations. Mutation events unrelated to integration have been found in both normal and REMI transformation. In some cases 50% of the mutations have not been tagged. Where restriction enzymes were used it is unclear if they stimulate these extra mutations. Improper repair of restricted genomic DNA canobviously lead to mutations without the insertion of transforming vector. The existence of untagged mutations make it critical to prove that the integrated DNA caused the mutation. Segregation analysis or gene disruption using the rescued plasmid from the integration event should be used to verify tagging before performing additional experiments with a mutant. Untagged mutants are still useful; gene cloning will just be more difficult.
Other considerations. Another important consideration is handling of primary transformants. Unless transformants are purified, workers risk masking valuable mutations due to heterokaryons formed during transformation. In this regard REMI mutagenesis is more time-consuming than UV mutagenesis of conidia where primary potential mutants require no manipulation before phenotype testing. Finally, it should go without saying that REMI is no substitute for a fundamentally sound transformation system with good selectable markers, high quality screens, and genomic and cDNA libraries necessary for gene cloning andanalysis. Gene tagging simply allows these tools to be used more efficiently.
Mutant Hunt Results
The largest REMI attempts have been undertaken in M. grisea where three different labs (7, 10, 11, 12) have tagged more than 30 genes relating to pathgencity and conidiation. At least a dozen of these genes have been cloned. Large efforts have also been attempted in Ustilago maydis (6, 14) where genesrelating to pathogencity have been cloned and Cochliobolus heterostrophus (3,15) where the TOX1 locus has been tagged. Significant efforts are also underway for the fungi listed with the email addresses.
REMI tagging has accelerated the interest of researchers in using molecular genetics to explore filamentous fungi, especially in species that were little studied in the past. Though not a magic bullet, the stimulation in transformation rates and the simplification and perhaps randomization of integration events makes the use of REMI a promising technique for gene tagging.
Noon-7:00 pm Registration, Holiday Inn-Worlds Fair
7:00 Hertel Foundation Lecture.
Convener: Barry Rolfe. Speaker: Fred Ausubel
8:30 pm Reception, Convention Center Ballroom
8:30 am Session I: Signal Transduction.
Convener: Dan Klessig
8:35 am Overview Lecture: Thomas Boller
9:20 am Thorsten Nurnberger
9:45 am Chris Lamb
10:10 am Break
10:40 am Phil Low
11:05 am Greg Martin
11:30 am Jean-Pierre Metraux
12:10 pm Lunch
1:30 pm Session II: Plant Resistance.
Convener: (to be announced)
1:35 pm Overview Lecture: Jeff Dangl
2:20 pm Jonathan Jones
2:50 pm Brian Staskawicz
3:20 pm Barbara Baker
3:50 pm Break
4:20 pm Session IIIA: Receptors?
Convener: Wolfgang Knogge
4:25 pm Felice Cervone
4:50 pm Jurgen Ebel
5:15 pm Jeff Ellis
5:40 pm Marilynn Etzler
6:05 pm Naoto Shibuya
4:20 pm Session IIIB: Biocontrol.
Convener: (to be announced)
4:25 pm Linda Thomashow
4:50 pm Ben Lugtenberg
5:15 pm Jim Ligon
5:40 pm Jo Handelsman
6:05 pm Leland Pierson
8:30 am Session IV: Biocontrol Overview Lecture.
Convener: Bonnie Ownley
8:35 am Joyce Loper
9:20 am Session V: Diversity and Ecology of Plant-Associated
Convener: Mariangela Hungria
9:25 am Steve Lindow
9:50 am Frans de Bruijn
10:15 am Break
10:45 am Chris Schardl
11:10 am Don Phillips
11:35 am Esperanza Martinez-Romero
12:10 pm Lunch
1:30 pm General Lecture I.
Convener: Jan Kijne.
Speaker: Ted Farmer
2:30 pm Session VI: Agrobacterium.
Convener: Mary Dell-Chilton
2:35 pm Overview Lecture: Eugene Nester
3:20 pm Stan Gelvin
3:45 pm Barbara Hohn
4:10 pm Peter Christie
4:35 pm Paul Bundock
5:00-7:00 pm Poster Session I
7:00 pm Southern Hospitality Barbecue-Worlds Fair Park
8:30 am Session VII: Bacterial Determinants for
Pathogenicity and Avirulence.
Convener: Mike Daniels
8:35 am Overview Lecture: Noel Keen
9:20 am Carol Bender
9:45 am Janet Leach
10:10 am Break
10:40 am Steve Farrand
11:05 am Christian Boucher
11:30 am Ulla Bonas
Free Afternoon for Tours and Activities
8:00 am Session VIII: Plant-Microbe Symbioses.
Convener: Ton Bisseling
8:05 am Overview Lecture: Sharon Long
8:50 am Herman Spaink
9:15 am Jens Stougard
9:40 am Katharina Pawlowski
10:10 am Break
10:40 am Adam Kondorosi
11:05 am Jean Denarie
11:30 am Birgitta Bergman
12:10 pm Lunch
1:30 pm General Lecture II
Convener: (to be announced).
Speaker: (to be announced)
2:30 pm Session IXA. Secreted Proteins.
Convener: Arun Chatterjee
2:35 pm George Salmond
3:00 pm Alan Collmer
3:25 pm Break
3:45 pm Dean Gabriel
4:10 pm Steve Hutcheson
4:35 pm Bill Broughton
5:00 pm Allan Downie
5:25 pm Peter Mueller
2:30 pm Session IXB: Emerging Areas.
Convener: A. B. Legocki
2:35 pm Mike Sadowsky
3:00 pm Martha Hawes
3:25 pm Break
3:45 pm Charles Opperman
4:10 pm Speaker: (to be announced)
4:35 pm Maria Harrison
5:00-7:00 pm Poster Session II
8:30 am Session X: Genetics of Fungal Pathogenicity.
Convener: Sally Leong
8:35 am Overview Lecture: Barbara Valent
9:20 am Regine Kahman
9:45 am Pierre de Wit
10:10 am Break
10:40 am Donald Nuss
11:05 am Jonathan Walton
11:35 am Anne Osbourn
12:10 pm Lunch
1:30 pm Session XIA: Molecular Virology.
Convener: David Hacker
1:35 pm Paul Ahlquist
2:00 pm Ichiro Uyeda
2:25 pm Linda Hanley-Bowdin
2:50 pm Break
3:20 pm Jim Culver
3:45 pm Bill Lucas
1:30 pm Session XIB: Rhizobium Signals.
Convener: Alfred Puhler
1:35 pm Brad Reuhs
2:00 pm Graham Walker
2:25 pm Michael Djordjevic
2:50 pm Break
3:20 pm Dale Noel
3:45 pm Dan Roberts
4:10 pm Session XII. Biotechnology.
Convener: Peter Gresshoff
4:15 pm Roger Beachy
4:40 pm John Ryals
5:05 pm Speaker: (to be announced)
5:40 pm Meeting Summary.
Convener: Evan Roth.
Speaker: Jim Cook
6:10 pm Close
7:00 pm Banquet, Convention Center Ballroom
IS-MPMI is an international society engaged in research in the areas of plant-microbe interactions and plant molecular biology. With the increase in member growth the Board wants to create an image for the society through the development of a logo. The following logo renditions incorporate four elements the board views as important to this image: global, molecular, fungi and plant pathology. Now we would like to hear from you. Return this form to Corie Dacus at IS-MPMI headquarters indicating your favorite logo or e-mail your comments to email@example.com, fax (651) 454-0766.
The proceedings from the 1996 8th International Congress on Molecular Plant-Microbe Interactions will be made available as the first book published with IS-MPMI's own imprint. Meeting registrants will receive a copy of this book as part of their registration sometime after the meeting. Books will also be available for sale to those who are unable to attend the Congress. This hardbound 6" x 9" book will be approximately 600 pages and will include information from more than 80 invited speakers. Watch for more information in upcoming issues of the IS-MPMI Reporter.
May 25-29, 1996
International Symposium on Bacterial Genetics and Ecology
Nauplion, Greece. Contact Ar. Amalia Karagouni, Institute of General Botany, University of Athens, Athens 15781 Greece. Fax: 301 7234136
June 11-14, 1996
Agricultural Biotechnology International Conference (ABIC)
Saskatoon, Saskatchewan, Canada. Contact: Ag-West Biotech Inc., 230-111 Research Drive, Saskatoon, SK S7N 3R2, Canada. Phone: 306/975-1939 Fax: 306/975-1966E-mail: firstname.lastname@example.org
June 16-21, 1996
Gordon Research Conference on Cellular and Molecular Mycology
Holderness School, Plymouth, New Hampshire, US. Contact: Dr. Anne Desjardins, Phone: 309/681-6378 Fax: 309/681-6686 E-mail: email@example.com
June 23-28, 1996
11th International Botrytis Symposium
Wageningen, Netherlands. Contact: Dr. Jan van Kan, Wageningen Agricultural Univ., Phone: 31 8370 83126 Fax: 31 8370 83412 E-mail: firstname.lastname@example.org
July 7-12, 1996
The Third International Nematology Congress
Gosier, Guadeloupe (French West Indies), Contact: Dr Alain Kermarrec, INRA-URPV, Local Arrangement Committee, BP 515, 97165, Pointe-a-Pitre cedex, Guadeloupe (FWI) Fax: (590) 94 11 72 / 25 59 02 Tel: (590) 25 59 40 E-mail: email@example.com
July 14-19, 1996
Eighth International Molecular Plant-Microbe Interactions Congress
Knoxville, Contact: UT Conferences, The University of Tennessee, P.O. Box 2648, Knoxville, TN 37901 USA Phone: 615/974-0250, Fax: 615/974-0264, E-Mail: firstname.lastname@example.org http://www.ce.utk.edu/departments/conferences/confhome.html
July 27-31, 1996
American Phytopathological Society Annual Meeting
Indianapolis, Indiana, U.S. Contact: Deb Merritt, Phone: (651) 454-7250 Fax: (651) 454-0766 E-mail: email@example.com
July 27-31, 1996
"Plant Biology '96" the 1996 American Society of Plant Physiologists Annual Meeting
Contact: Susan Chambers, ASPP headquarters, 15501 Monona Drive,Rockville, MD 20855-2786 USA Phone: 301-251-0560 ext 11 Fax: 301-279-2996, E-mail: firstname.lastname@example.org
July 30-August 8, 1996
5th International Oat Conference and 7th International Barley Genetics
Univ. of Saskatchewan, Saskatoon, Saskatchewan, Canada. Contact: C. Ouellet, Univ. of Saskatchewan, Fax: 306/966-5015
August 4-9 1996
First International Conference on Mycorrhizae (ICOM 1)
Contact: Tom Bruns Department of Environmental Science, Policy and Management 108 Hilgard Hall, University of California, Berkeley, California 94720-3110, USA
August 11-16, 1996
Queenstown Molecular Biology Meeting
Queenstown, New Zealand. Contact: Billie Masters, PO Box 653, Dunedin, New Zealand. Fax 64 3 477 7757
August 25-29, 1996
Sixth International Fungal Spore Conference
University Konstanz, Germany. Contact: Kurt Mendgen, Universitat Konstanz, Postfach 5560, D-78434, Konstanz, Germany. Fax 49-7531-883035 Phone: 49-7531-883680
September 1-5, 1996
Symposium on Plasmid Biology (Plasmid Biology '96)
Contact: Helmut Schwab, Institut f. Biotechnologie, Technische University of Graz, A8010 Graz, Austria. Fax: 43-316 873-8434
September 12-15, 1996
Fallen Leaf Lake Conference on Horizontal Gene Transfer: Implications and Consequences
Leaf Lake, South Lake Tahoe, California, U.S. Contact: Michael Syvanen, Dept. Of Microbiology & Immunology, University of California, Davis, CA 95616. E-mail: email@example.com
March 18-23, 1997
The 18th Fungal Genetics Meeting
Asilomar, California, U.S., Contact: Dr. N. Louise Glass, Biotechnology Laboratory, University of British Columbia, Vancouver, BC V6T 1W5 Canada Fax: 604/822-6097 or Dr. Michael J. Hynes, Department of Genetics, University of Melbourne, Parkville, VIC 3052 Australia, Fax: 613/934-45139 E-mail: firstname.lastname@example.org http://www.kumc.edu/*fgsc
June 22-27, 1997
The Second International Bacterial Wilt Symposium
Gosier, Guadeloupe (French West Indies) Contact: Dr. Philippe, Prior, INRA, B.P. 515, 97165 Pointe-a-Pitre, Guadeloupe, French West Indies, Fax: (590)94 11 72 E-mail: email@example.com
If you have a meeting you would like to list here, please send
the information to:
Corie Dacus, IS-MPMI, 3340 Pilot Knob Road, St. Paul, MN 55121-2097
Phone: (651) 454-7250, Fax: (651) 454-0766, E-mail: firstname.lastname@example.org
This year's International Molecular Plant-Microbe Interactions Congress plans to be bigger than ever. The scope is impressive with joint sessions of interest to microbiologists, geneticists, pathologists, chemists, agronomists, and others.
These meetings are held only once every two years so we hope you don't miss this opportunity to interact with fellow scientists and learn of significant advances in our field.
As Eugene Nester mentioned in his President's Column in this issue, our Society is growing and the needs of our members are increasing. Members have told us that they have a greater interest in staying informed on the Society's activities and in finding ways to interact with other members.
One of the ways to do this is through the IS-MPMI Reporter. Let fellow members know of upcoming meetings, research developments, technological advancements, etc. Below is a schedule of the next two issues of IS-MPMI Reporter for your reference. In the meantime, we hope to see you in Tennessee.
Summer issue (includes member directory and post
Articles due: August 1
Mail date: August 30
Articles due: December 1
Mail due: December 30
Plant Disease Unit
CNRS Institut de Sciences Vegetales
Eugene W. Nester
Dept. of Microbiology, SC-42
University of Washington
Seattle, WA 98195 USA
Barry G. Rolfe
Australian National University
GPO Box 475Canberra ACT 2601
U.S. Dept. of Agriculture - ARS
Washington State University
Pullman, WA 99164-6430 USA
U.S. Dept. of Agriculture - ARS
University of Wisconsin
Madison, WI 53706 USA
Steven C. Nelson
3340 Pilot Knob Road
St. Paul, MN 55121 USA