In late 2017, we sampled host-seeking I. scapularis ticks at 70 forested sites in 26 low-elevation counties in the upper Tennessee Valley (Figure 1, panel B). To find tick habitats (hardwood or conifer forests <800 m in elevation) accessible for sampling (i.e., trails through public forests or margins of public roads through private forests), we reviewed Google Earth (https://www.google.com/earth/) satellite imagery. We sampled each site once during the peak of adult I. scapularis tick activity (late October–January). We recorded site elevation and geo-coordinates and collected host-seeking ticks using a standardized drag-cloth method; in brief, we dragged a 1-m² white corduroy cloth across leaf litter and checked every 10 paces for attached ticks. We dragged cloths 30–60 minutes per site and described tick tallies as number collected per hour to correct for variations in effort per site. We did not conduct drag-cloth collections during periods of rain, strong wind, low air temperatures (<8°C), or low relative humidity (<40%).

We placed ticks in 70% ethanol, identified species using a morphologic key (6), and tested ticks for Borrelia spirochete infection by DNA extraction and quantitative multiplex real-time PCR using differential probes targeting the 16S rDNA of Lyme group Borrelia and relapsing fever group Borrelia (7). We then subjected a random subset of negative samples and samples positive by the 16S assay (maximum 6 samples/site) to PCR amplification of the 16S–23S rDNA intergenic spacer region (8) and Sanger sequencing for species-level identification.

No previous tick drag-cloth counts existed for the counties in our survey area, except for a 1,050-m transect of land in a forest in Anderson County, which we have drag-cloth sampled annually each December since 2012. To assess a trend in adult I. scapularis tick abundance, we applied linear regression modeling to the tick tallies from that transect of land.

In late 2017, we collected 479 adult I. scapularis ticks from 49 of 70 sites in the upper Tennessee Valley. Two adult Amblyomma americanum ticks collected during the survey were excluded from analysis. We detected I. scapularis ticks in all 26 counties surveyed, 23 of which met the criterion used by Eisen et al. for established I. scapularis populations (Figure 1, panel B) (3). Site elevations were 210–730 m; the highest elevation at which I. scapularis ticks were found was 570 m. The average number of adult ticks collected per hour during drag-cloth surveys was 8.8 (range 0–48). At the Anderson County site that had been drag-cloth sampled annually, a highly significant increasing trend in I. scapularis ticks was evident (p = 0.003; Figure 2); the count in 2017 (24.8 ticks/hour) was 3.5× higher than that in 2012.

We tested all I. scapularis ticks collected (N = 479) for Borrelia spp. infection; 46 ticks (9.6%) from 7 sites in 4 counties (Anderson, Claiborne, Hamilton, and Union; Figure 1, panel B) tested positive for Lyme group Borrelia by 16S rDNA PCR screening. We tested 26 samples for the intergenic spacer region by PCR; all were positive for this sequence and identified as B. burgdorferi sensu stricto by sequencing. Most infected ticks came from 2 Union County sites, which had prevalences of 44% (14/32) and 78% (18/23). No ticks were found to be infected with B. miyamotoi or other relapsing fever group borreliae.

In eastern Tennessee, public awareness and concern about ticks focuses primarily on the abundant lone star ticks (Amblyomma americanum) and American dog ticks (Dermacentor variabilis) encountered during the spring and summer. Both species can spread pathogens (9), but neither are vectors of B. burgdorferi spirochetes. Immature I. scapularis ticks are similarly active in the summer, but in southern states, these ticks typically avoid host-seeking above leaf litter and are rarely seen on humans or drag-cloths (10). For this reason, assessment of I. scapularis distribution in southern states is best achieved by acquiring adult life-stage ticks during cool season drag-cloth surveys (as reported here) or by collecting ticks from deer harvested in the fall. Inspection of hunter-harvested deer is efficient for the detection of low-density I. scapularis ticks (11). Thus, our drag-cloth sampling for I. scapularis ticks in 14 counties where none were found on deer a decade ago (Figure 1, panels A, B) suggests that tick abundance in these counties has increased. This suggestion is supported by a >3-fold increase in I. scapularis tick counts at the Anderson County site where we have 6 consecutive years of drag-cloth counts.

This study documents emergence of B. burgdorferi senso stricto in tick populations in eastern Tennessee. Infected ticks were predominantly found in high-prevalence hot spots in Union County (36.39°N). Relative to Lyme disease–endemic areas in the north, B. burgdorferi prevalence in the study area was low (10%) and had a patchy distribution (7/49 sites had positive ticks). This distribution could reflect host barriers of B. burgdorferi transmission in the South (12), or more concerning, the hot spots in Union County might reflect the beginning of an infection surge, similar to that seen in southwestern Virginia during the past decade (4).

In the United States, Lyme disease is primarily a summertime disease associated with bites from nymphal I. scapularis ticks. In southern states, detection of B. burgdorferi bacteria in adult ticks does not necessarily imply risk to humans; for example, B. burgdorferi cycles in I. scapularis populations on the Outer Banks of North Carolina, yet nymphs in that area cannot be collected on drag-cloths and no locally acquired cases of Lyme disease have been reported (13). In contrast, infected nymphs have been found on drag-cloths from surveys in Virginia, where Lyme disease incidence has spiked (14). We speculate that Borrelia-infected I. scapularis populations emerging in southwestern Virginia include immigrant ticks from the North, with some nymphs in these populations exhibiting host-seeking behaviors that lead to contact with humans. A similar invasion process might be under way in eastern Tennessee; the surveillance data reported here provide a baseline for investigating this possibility. Health officials and practitioners need to be vigilant for increasing Lyme disease incidence in Tennessee.