LYME DISEASE RISK ASSESSMENT, FORT LEONARD WOOD, MISSOURI, 26 APRIL-2 MAY 1992 DEPARTMENT OF THE ARMY US Army Environmental Hygiene Activity-West Fitzsimons Army Medical Center Aurora, Colorado 80045-5001 [Seal of Department of Defense, United States of America] REPLY TO ATTENTION OF: HSHB-AW-P (40-5f) 24 July 1992 LYME DISEASE RISK ASSESSMENT NO. 16-66-AI44-92 FORT LEONARD WOOD, MISSOURI 26 APRIL-2 MAY 1992 1. REFERENCES. See Appendix A for a list of references. 2. AUTHORITY. a. Conference Report on the National Defense Authorization Act for Fiscal Years 92 and 93, House Report 102-311, 102nd Congress, 1st Session, 13 November 1991, p. 422. b. USAEHA Form 250-R, U.S. Army Health Services Command, 24 March 1992. 3. PURPOSE. The purpose in performing this risk assessment was to survey for Lyme disease vectors, to determine the prevalence of the infective agent, Borrelia burgdorferi, in rodents and ticks, and to assess the risk of Lyme disease to Fort Leonard Wood personnel. See Appendix B for other assistance that can be obtained from the U.S. Army Environmental Hygiene Activity-West (USAEHA-W). 4. GENERAL. a. Risk Definition. The term "risk", as used in this report, is a non- statistical determination of qualitative and quantitative information available to evaluate the potential to acquire Lyme disease. To the extent available, information evaluated includes the following elements of the Lyme disease cycle: (1) History of Lyme disease in the area. (2) The presence or absence of the tick vector (Ixodes scapularis) and the mammalian host population needed to sustain a viable population of the vector. (3) The presence of the Lyme disease-causing spirochete (B. burgdorferi) in the tick population or in the mammalian reservoir host population. (4) The presence of antibodies to B. burgdorferi in the mammalian host population. Criteria for risk categorization follow: Low risk - Some element of the Lyme disease cycle identified in nearby areas but not on the installation. Moderate risk - Elements of Lyme disease cycle identified from the installation or human cases of Lyme disease identified from the local area. High risk - All elements of the Lyme disease cycle are present on the installation. b. Dr. Lester D. Hale, the Survey Officer and SPC Reginald Flemming, a Preventive Medicine Specialist, discussed major findings and recommendations with Fort Leonard Wood personnel. A list of personnel contacted and personnel assisting Dr. Hale during the risk assessment is found in Appendix C. c. The field survey portion of the Lyme Disease study was conducted at Fort Leonard Wood on 26 April-2 May 1992. d. This study also provided training for Fort Leonard Wood Preventive Medicine personnel. On-the-job training conducted during the field survey portion of the study included: vector surveillance procedures, tick surveillance methods, small mammal trapping, and ear tissue biopsy procedures. Fort Leonard Wood personnel were exceptionally receptive to information on Lyme disease and personal protective measures for prevention of tick bites. e. Laboratory analyses of ticks and rodent ear biopsies were conducted between 29 April-16 July 1992. 5. BACKGROUND. a. Lyme disease is a multi-systemic infectious disease caused by the spirochete, B. burgdorferi, which is transmitted to humans by the bite of an infected tick. The disease is most often referred to as Lyme disease or Lyme arthritis in the United States. Lyme disease has become the most prevalent arthropod-borne illness in North America. Its geographic range is expanding and the number of reported cases continues to rise each year. The Office of the Surgeon General reported 379 cases of Lyme disease contracted on Department of Defense (DOD) installations from 1987 through 1991. During 1990, there were 78 cases of Lyme disease treated in military hospitals from all the Armed Services of which 26 of the cases were either dependents or retired members. During 1991, there were 81 cases of Lyme disease treated in military hospitals of which 31 cases were either dependents or retired members. There were 3 soldiers who were reported to have permanent disabilities caused by Lyme disease during 1991. The need to protect soldiers and other personnel working on DOD installations has increased with the spread of Lyme disease. b. According to the Centers for Disease Control (CDC), Fort Collins, Colorado (Reference 1, Appendix A), there were 193 cases of Lyme disease occurring in Missouri during 1991. There was one case of Lyme disease reported from Pulaski County in 1990. Preventive Medicine personnel stated that at the General Leonard Wood Community Hospital, physicians diagnosed two people with Lyme disease per year during 1990 and 1991, and one person in 1992. An epidemiology investigation indicated that these cases probably originated on Fort Leonard Wood. c. Laboratory analysis performed by this Activity on ticks collected at Fort Leonard Wood during the 1991 fall deer hunt showed that 40 of 112 (36 percent) western black-legged ticks, I. scapularis examined by immunofluorescent staining were positive for spirochetes (Reference 2, Appendix A). Ticks collected in 1987 at Fort Leonard Wood were negative for spirochetes. d. Fort Leonard Wood is located south of Interstate Highway 44 and approximately 72 miles northeast of Springfield in Pulaski County. St. Roberts is located about 2 miles north of the main gate entrance to the installation. The installation currently encompasses approximately 62,910 acres. The actual maneuver/training areas were approximately 41,918 acres. During 1991, Fort Leonard Wood provided initial entry training and professional training for approximately 78,192 Active troops, National Guardsmen and Reservists. In addition, weekend training was provided to 11,952 National Guardsmen and Reservists. 6. METHODS. a. Field Survey Procedures. (1) Tick Surveys. The tick collecting methods at Fort Leonard Wood included using tick drag cloths and/or tick flags, and carbon dioxide traps. Ticks were also removed from the ears of trapped rodents. All ticks were kept alive by placing them in water moistened vials as described in the USAEHA-W, Entomological Sciences Division (ESD) Standing Operating Procedures (SOP) No. 6 and No. 8. For each collection site, all collected ticks were identified, counted, and placed into a marked vial. Ticks collected at different sites were kept segregated and placed into separate marked vials and subsequently sent to the USAEHA-W, ESD Laboratory. (a) Tick Drag Technique. Two narrow boards (1x2x36-inch) were attached on opposite ends of a 3x3-foot white flannel cloth. A cord was fastened to the ends of one board. The cloth was then pulled slowly over vegetation for approximately 10 meters before examination for attached ticks. Ticks were then removed from the cloth and placed in a marked vial for that particular dragging operation. Approximately 9,750 linear meters were covered by team members at the different collection sites. (b) Tick Flag Technique. A 3x3-foot piece of white flannel cloth was dragged over the tops of large vegetation, i.e., sagebrush, or was used on steep slopes. Attached ticks were then removed from the cloth and placed in a marked vial for each collection site. Aproximately 3,250 linear meters were covered by team members doing the flagging. (c) Carbon Dioxide Traps. A 2x2-foot piece of white flannel cloth was placed on the ground and the edges were weighted down with stones. An aluminum-foil covered pie plate was placed upside down in the middle of the cloth. Approximately 2 pounds of dry ice were wrapped in newspaper and placed on top of the pan. The area around each trap and the cloth were examined for ticks after 3 to 4 hours. (2) Rodent Surveys. Rodents trapped at Fort Leonard Wood were identified using "A Field Guide to the Mammals, North America, north of Mexico" (Reference 5, Appendix A). All rodents were handled in accordance with the guidelines outlined in the National Institutes of Health Publication No. 85-23, Revised 1985, "Guide for the Care and Use of Laboratory Animals" and USAEHA-W, ESD SOP No. 7. (a) Small mammals were captured in live traps (3x3x10-inches) on Fort Leonard Wood. At each site several trapping locations were selected and one or more trap lines were set in each area. The traps were usually located near stream beds, springs, ponds, marshes, or rivers with traps located approximately 10 meters apart. The traps were set only one night at each location using at least 25 traps at each site at Fort Leonard Wood. (b) Each rodent was anesthetized before removing any ticks or taking an ear biopsy. The ear biopsy was placed in a labelled vial containing a phosphate buffer solution and then frozen on dry ice. All biopsies were kept frozen until assayed for Lyme disease. (3) Specimen Handling Procedures. The ticks collected from rodents and from dragging/flagging vegetation during 26-29 April were shipped via overnight mail to USAEHA-W, ESD Laboratory for bioassay and maintained at 18 degrees C until assayed for Lyme disease. All the ear tissue biopsies from Fort Leonard Wood and those ticks collected from 30 April-2May were hand-carried to the USAEHA-W, ESD Laboratory for Lyme disease bioassay. b. Laboratory Assay Procedures. (1) The midguts of 3 black-legged ticks, I. scapularis, 203 Lone Star ticks, Amblyomma americanum, and 19 American dog ticks, Dermacentor variabilis, were dissected and examined for spirochetes using immunofluorescent microscopy. Another 29 Lone Star ticks and 2 American dog ticks were dissected and examined for spirochetes using darkfield microscopy. (2) Genomic deoxyribonucleic acid (DNA) was extracted and amplified from A. americanum and D. variabilis using Polymerase Chain Reaction (PCR) by personnel at the CDC. Ticks were extracted in pools of 4 to 14 adults or 3 to 7 nymphs. Twelve pools of A. americanum and one pool D. variabilis were assayed for B. burgdorferi by the PCR technique. (3) Twenty-six A. americanum were analyzed using an Enzyme Linked Immunosorbent Assay (ELISA) for B. burgdorferi developed by the CDC. (4) Forty-nine rodent ear biopsies were cultured in BSK II media. Each inoculated culture was examined under the darkfield microscope every 3-7 days for 21 days for spirochete growth. If no spirochetes were found during this period, the samples were considered to be negative for B. burgdorferi. 7. FINDINGS AND DISCUSSION. a. Tick Collections. Ticks were collected from humid or wet sites such as stream banks, hillsides, and near a lake. Table 1 shows the sites, map coordinates, species, and number of each of the life stages of I. scapularis, A. americanum, and D. variabilis, collected from drags/flags and from carbon dioxide traps at Fort Leonard Wood. There were 3 I. scapularis, 27 D. variabilis, and 1,409 A. americanum collected by the various methods. Table 2 shows the number of ticks removed from rodents at Fort Leonard Wood. Amblyomma americanum was the only tick species collected from these rodents. b. Rodent Collections. Three species of rodents were captured at seven sites on Fort Leonard Wood (Table 2). The deer mouse was the most common rodent captured. c. Laboratory Results. Detailed information on the sites, species of ticks and rodents, and laboratory results are given in Appendix D. TABLE 1. Number of Ixodes scapularis, Amblyomma americanum, and Dermacentor variabilis, collected from drags/flags and from carbon dioxide traps at Fort Leonard Wood, Missouri. NUMBER OF TICKS -------------------------------------------- SITE I. scapularis A. americanum D. variabilis --------------------------------------------------------------------------- East Gate Road Creek 1A 29A,15N 1A (814792)* Happy Hallow Rec Area 0 19A,5N 0 (765838) Boy Scout Camp 0 16A,9N 0 (832788) Big Piney Camp Grounds 0 51A,33N 0 (826796) Training Area 236 0 9A,3N 2A (735719) Range 25 0 36A,6N 8A (744709) Bloodland Lake 0 57A,103N 13A (742715) Hunting Area 33 0 84A,97N 1A (743805) Training Area 204N 2A 64A,198N 1A (705746) Piney Valley Golf Course 0 18A,17N 1A (816744) West of Training Area 244 0 126A,269N 0 (728755) Road by Range 20, 21, and 22 0 39A,36N 0 (747664) Wood Middle School and Housing 0 51A,19N 0 (767816) Stream Behind Service (Did not drag/flag in this area; did trap 2 mice) Station (776817) ---------------------------------------------------- TOTAL TICKS COLLECTED 3A 599A,810N 27A * Map coordinates given as longitude and latitude; based on Fort Leonard Wood Reservation Special Map (Missouri 1:50,000), Published by the 63rd Engineer Company, dated 1979. A - Adult N - Nymph -------------------------------------------------------------------------- (1) Ticks. All of the ticks examined by darkfield and immunofluorescent microscopy were negative for B. burgdorferi. In addition, all tick pools or individual ticks tested by the PCR and the ELISA techniques were negative for this spirochete. (2) Rodent Ear Biopsies. Forty-nine rodent ear biopsies were cultured in BSK II media for spirochetes (Table 2). All of these samples were negative for B. burgdorferi, suggesting that the spirochete had not circulated within the limited population sampled at Fort Leonard Wood. TABLE 2. Number of Amblyomma americanum removed from rodents at Fort Leonard Wood, Missouri. RODENTS ------------------- SITE No. Species A. americanum --------------------------------------------------------------------------- East Gate Road Creek 3 Peromyscus maniculatus 29L,6N (deer mouse) Boy Scout Camp 4 Peromyscus leucopus 1L (white-footed mouse 6 P. maniculatus 34L 1 Microtus ochrogaster 0 (prairie vole) Big Piney Camp Grounds 3 P. leucopus 2L 1 P. maniculatus 0 Bloodland Lake 4 P. leucopus 26L 9 P. maniculatus 32L,2A 1 M. ochrogaster 0 Hunting Area 33 4 P. leucopus 17L,2N 6 P. maniculatus 31L,3N West of Training 5 P. maniculatus 32L Area 244 Stream Behind Service 1 P. leucopus 2L Station 1 P. maniculatus 3L,1N ----- ---------- TOTALS 49 209L,14N L - Larva, N - Nymph, A - Adult --------------------------------------------------------------------------- d. Lyme Disease Risk. (1) During the field survey portion of this Lyme Disease Risk Assessment, many ticks were collected by dragging/flagging, from carbon dioxide traps, or removed from trapped rodents. This information reflected the comments from installation personnel that people collect ticks on themselves when outdoors on Fort Leonard Wood. (2) One black-legged adult tick was found during dragging/flagging at the East Gate Road Creek site and two adult ticks were found during dragging/flagging at Training Area 204N site. This species is a proven vector of Lyme disease and has the potential to acquire and transmit this disease. Many Lone Star ticks were collected from dragging/flagging, from carbon dioxide traps, and from captured rodents at Fort Leonard Wood. The Lone Star tick has been suspected as a vector of Lyme disease; however, no spirochetes were detected from the samples collected from this species. Twenty-seven American dog ticks were collected; however, this tick species has not been implicated as a vector of Lyme disease. (3) No Lyme disease spirochetes were recovered from either the ticks or from the rodent ear tissue samples collected during this study. However, the presumption at this time, based on this Activity's previous collected data (Reference 2, Appendix A), is that spirochetes were present in the black-legged ticks collected from deer. (4) To determine the human health risk from Lyme disease at Fort Leonard Wood, the Risk Definition as listed in paragraph 4a, this report was used. The data collected during this assessment and prior studies done on ticks collected from Fort Leonard Wood indicate that all the criteria for a High risk for acquiring Lyme disease are present. Human cases have been documented as originating on the installation. Although the criteria suggest a High risk, the actual risk to troops training in the field is probably the lower Moderate risk based on the relative low population density of I. scapularis encountered during the assessment. Personal protective measures to guard against tick bites would be prudent. Risk of contracting Lyme disease can be minimized by the proper wearing of clothing, by avoiding areas known to harbor high tick populations, and by the use of repellents (See Appendix E for repellent products available in the Defense General Supply Center or Self Service System). General Lyme disease reduction measures can be found in Appendix F. (5) This Activity had coordinated collection of ticks from Fort Leonard Wood during the fall 1991 deer hunts. Ticks sent to our USAEHA-W, ESD Laboratory were identified and analyzed for Lyme disease. Continuing studies such as these should further define the presence or absence of Lyme disease. 8. CONCLUSIONS. The black-legged tick, I. scapularis, the tick vector species for Lyme disease in Missouri, was collected at Fort Leonard Wood. Borrelia burgdorferi, the causative agent for Lyme disease, was not found in the tick and small mammal populations sampled during this assessment. All the criteria for a High risk for acquiring Lyme disease were present on Fort Leonard Wood. Even though the criteria suggest a High risk, the actual risk to troops training in the field is probably a Moderate risk based on the relative low population density of I. scapularis encountered during this assessment. However, if populations of I. scapularis were to increase at Fort Leonard Wood, then the risk to troops would be High. [signature] LESTER D. HALE Entomologist APPROVED BY: [signature] RONALD J. RAKICKAS LTC, MS Chief, Entomological Sciences Division APPENDIX A REFERENCES 1. Lyme Disease Surveillance Summary, Volume 3, No. 1, March 1992, Centers for Disease Control. 2. Memorandum, this Activity, HSHB-AW-P, 22 January 1992, subject: Lyme Disease Risk Assessment, Project No. 16-66-AF35-92, Fort Leonard Wood, Missouri. 3. USAEHA-W, ESD SOP No. 6, 24 April 1991, Tick Collection Procedures. 4. USAEHA-W, ESD SOP No. 8, 25 April 1991, Small Mammal Tick and Ear Tissue Collections. 5. Burt, W. H. and R. P Grossenheider, A Field Guide to the Mammals North America north of Mexico, Third Edition, 1976, Houghton Mifflin Company, Boston, 289pp. 6. National Institutes of Health Publication No. 85-23, Revised 1985, Guide for the Care and Use of Laboratory Animals, U.S. Department of Health and Human Services. 7. USAEHA-W, ESD SOP No. 7, 25 April 1991, Procedures for Trapping and Handling Small Mammals. APPENDIX B TECHNICAL ASSISTANCE Technical advice and/or consultation on pest management problems, to include on-site assistance, may be obtained by telephone from our Activity, DSN 943-8090. Please inform your Major Command Pest Management Consultant if you desire to request on-site assistance from our Activity. Technical services that we can assist you with are as follows: 1. Entomological laboratory support 2. Environmental laboratory support 3. Hazardous waste management 4. Industrial hygiene management 5. Medical systems safety and health 6. Sanitation and hygiene 7. Wastewater management 8. Water supply management 9. Worksite hazards management 10. Cholinesterase testing management For assistance in any of the above listed programs, please call: Environmental Health and Engineering Division - DSN 943-8100 Field sanitation and hygiene; potable, recreational and wastewater quality; hazardous waste management; document/design reviews. Industrial Hygiene Division - DSN 943-8881 Industrial hygiene; hazard communication; protective equipment programs; document/design reviews. Environmental Laboratory Division - DSN 943-3293 Routine and emergency analysis of water, soil, and occupational health- related samples. Cholinesterase Laboratory Division - DSN 943-4838 Testing of red blood cell-cholinesterase (RBC-ChE) specimens and quality assurance consultations and training for RBC-ChE labs. During non-duty hours calls will be recorded by an answering machine and returned the next day. Many additional services are available from our parent organization, the U.S. Army Environmental Hygiene Agency, and are described in AEHA Pamphlet 40-2, Directory of Services (published annually). We will gladly coordinate any additional services you request and that we cannot provide, with our parent organization. APPENDIX C PERSONNEL CONTACTED 1. COL Thomas H. Auer (*) Hospital Commander DSN 581-9131 2. LTC Troy W. Bradshaw (*) Chief, Preventive Medicine Service DSN 581-1061 3. Mr. Donnie Gorman (*H) Environmental Health Technician DSN 581-1063 4. SPC Phoebe E. Long (H) Preventive Medicine Specialist DSN 581-1062 5. SPC Jay Reed (H) Preventive Medicine Specialist DSN 581-1062 6. SPC Magda Serrano (H) Preventive Medicine Specialist DSN 581-1062 ----------- * Individual received an inbriefing. H Individual assisted with the field portion of the Lyme disease risk assessment. APPENDIX D TICK LOG 1992 [Data omitted - data summarized in above report] APPENDIX E REPELLENTS 1. Several arthropod repellents are available through the Defense General Supply Center (DGSC) or Self Service Supply System. When used in accordance with directions on the label and in conjunction with the proper wearing of clothing, they provide personal protection against a wide variety of medically important insect/arthropod pests. Availability and current pricing can be obtained by calling the DGSC at DSN 695-4865 or commercial (804) 790-4865. Repellents available for use are described below: a. Insect/Arthropod Repellent Lotion (cream, 2 fluid ounces) for application to exposed skin. The lotion, NSN 6840-01-284-3982, is not labeled for ticks, but will repel chigger mites and many biting flies. b. Permethrin Arthropod Repellent, Insect Repellent, Clothing Application (aerosol, 6 ounces) NSN 6840-01-278-1336. Seventy-five percent of the can is used to apply to the field uniform and the remainder is used to treat mosquito netting. The product provides protection from ticks and mosquitoes for a maximum of five weeks or five launderings. Apply more frequently if "buddy checks" reveal attached ticks. c. Insect Repellent Fabric Treatment (liquid, 5.1 fluid ounces) NSN 6840-01-334-2666. The contents are added to 2 gallons of water and applied with the 2-gallon sprayer from a field sanitation kit at a pressure of 55 pounds per square inch to field uniforms, mosquito netting, and tent fabric to provide protection from ticks, biting flies, and other insects. Since most sprayers are not equipped with the required pressure gauge (NSN 3740- 01-332-8746), it will be necessary to obtain a pressure gauge and filter (NSN 4330-01-332-1639), in order to complete the retrofitting. Proper application can provide protection for the normal life of the uniform (180 days in the field), six launderings of mosquito netting, and 6-9 months of treatment for tent fabric, depending on the climate. 2. Detailed directions for the use of these and other repellents can be found in the U.S. Army Environmental Hygiene Agency Technical Guide (TG) 174, Personal Protective Techniques Against Insects and Other Arthropods of Military Significance, June 1991. 3. The U.S. Army Medical Department Tick-Borne Disease Card (7189) is available from the USAEHA-W, ESD. APPENDIX F Lyme Disease Risk Reduction Measures 1. Emphasize public awareness programs to educate troops, dependents, civilian employees and visitors on personal protective measures and Lyme disease. Methods should include, but not be limited to: a. Distribution of printed Lyme disease handouts, such as tick identification cards (USAMD-7189), pamphlets, and fact sheets. b. Notifications in the installation newsletter and post electronic media (e.g., closed-circuit TV), especially prior to the high-risk months (April-September). c. Making available, for viewing, video and 35mm slide format presentations on Lyme disease that are available from this Activity. 2. Submit any collected tick specimens (both field-collected or ticks that have been removed from individuals) for identification and immunofluorescent staining or darkfield microscopy testing to the Entomological Sciences Division, U.S. Army Environmental Hygiene Activity-West, Fitzsimons Army Medical Center, Aurora, CO 80045-5001. 3. Stock Permethrin Arthropod Repellent (NSN 6940-01-278-1336, box of 12 cans for $36.99), and 3M [Trademark] Insect Repellent (NSN 6840-01-284- 3982, box of 12 tubes, $29.30) for distribution. Emphasize tick habitat avoidance and the proper wearing of clothing and use of repellents. 4. Report all confirmed and suspected cases of Lyme disease [e.g., suspicious febrile illnesses, arthralgias, rashes, (Erythema Migrans)] by special telegraphic report [MED-16(R4)] for all soldiers and civilian medical care beneficiaries. 5. Identify high risk foci in cantonment areas via tick dragging/flagging, small mammal trapping, deer checks, and the assaying of collected ticks for B. burgdorferi. Sampling should be performed in early summer when I. scapularis nymphs (the life stage responsible for most human Lyme disease infections) are active. Post DA Poster 40-5, and thereby identify high risk areas. DA Poster 40-5 can be obtained by writing to the Commander, U.S. Environmental Hygiene Agency, ATTN: HSHB-MR-E, Aberdeen Proving Ground, MD 21010-5422 or by telephone: DSN 584-3613 or Commercial (410) 671-3613. 6. Avoid high tick population areas for troop training or recreation. Such areas can be identified by tick dragging or flagging prior to use. Case by case surveillance is necessary due to the patchy distribution of I. scapularis. 7. Eliminate tick habitat in heavily used, infested areas (e.g., wooded recreation areas) by removing low brush and leaf litter. Tick infestations should be verified via tick flagging or dragging prior to habitat modification. Clearing should be done in low risk months (i.e., November - February). 8. Prepare, as a contingency, to treat high-use areas with pesticides to decrease tick numbers if surveillance reveals high tick numbers and if nonchemical control techniques (e.g., brush removal, mowing, raking) do not provide adequate control. --- [Trademark] 3M is a registered trademark of Minnesota Mining and Manufacturing Co., St. Paul, MN 55133-3053