Associations of passerine birds, rabbits, and ticks with Borrelia miyamotoi and Borrelia andersonii in Michigan, U.S.A

Abstract

Background: Wild birds contribute to maintenance and dissemination of vectors and microbes, including those that impact human, domestic animal, and wildlife health. Here we elucidate roles of wild passerine birds, eastern cottontail rabbits (Sylvilagus floridanus), and Ixodes dentatus ticks in enzootic cycles of two spirochetes, Borrelia miyamotoi and B. andersonii in a region of Michigan where the zoonotic pathogen B. burgdorferi co-circulates.

Methods: Over a four-year period, wild birds (n = 19,631) and rabbits (n = 20) were inspected for tick presence and ear tissue was obtained from rabbits. Samples were tested for Borrelia spirochetes using nested PCR of the 16S-23S rRNA intergenic spacer region (IGS) and bidirectional DNA sequencing. Natural xenodiagnosis was used to implicate wildlife reservoirs.

Results: Ixodes dentatus, a tick that specializes on birds and rabbits and rarely bites humans, was the most common tick found, comprising 86.5% of the 12,432 ticks collected in the study. The relapsing fever group spirochete B. miyamotoi was documented for the first time in ticks removed from wild birds (0.7% minimum infection prevalence; MIP, in I. dentatus), and included two IGS strains. The majority of B. miyamotoi-positive ticks were removed from Northern Cardinals (Cardinalis cardinalis). Borrelia andersonii infected ticks removed from birds (1.6% MIP), ticks removed from rabbits (5.3% MIP), and rabbit ear biopsies (5%) comprised twelve novel IGS strains. Six species of wild birds were implicated as reservoirs for B. andersonii. Frequency of I. dentatus larval and nymphal co-feeding on birds was ten times greater than expected by chance. The relatively well-studied ecology of I. scapularis and the Lyme disease pathogen provides a context for understanding how the phenology of bird ticks may impact B. miyamotoi and B. andersonii prevalence and host associations.

Conclusions: Given the current invasion of I. scapularis, a human biting species that serves as a bridge vector for Borrelia spirochetes, human exposure to B. miyamotoi and B. andersonii in this region may increase. The presence of these spirochetes underscores the ecological complexity within which Borrelia organisms are maintained and the need for diagnostic tests to differentiate among these organisms.

Figure 1

Field investigations of wild birds and eastern cottontails for the presence of ticks.A) American Robin harbors a single nymphal tick (near the tip of forceps); B) multiple engorged larval ticks are present beneath the auricular feathers and within the skin of the ear canal of a White-throated Sparrow; C) juvenile eastern cottontail harbors an engorged adult I. dentatus near the scruff; D) adult ticks of differing engorgement status are present on an eastern cottontail. Photo credits: Gabriel Hamer and Graham Hickling.

Figure 2

Phenology of larval and nymphal bird-associated ticks.A) Ixodes dentatus; B) H. leporispalustris depicted as weekly mean proportions of infested birds (error bars are standard error of the mean across years 2004–2007).

Figure 3

Neighbor-joining phylogenetic tree based on 476 nucleotides of B. miyamotoi 16S-23S rRNA IGS haplotypes collected from Pitsfield Banding Station, 2004–2007. A sequence of B. lonestari obtained from GenBank was included as the outgroup. The twelve sequences generated in the current study are in bold and comprised two strain types, including 11 sequences that are identical to ‘Type 4’ (accession no. GU993308) and one novel strain indicated by the red triangle (GU993309). The percentages of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) are shown next to the branches when 60 or higher. Sequences are labeled with the name of the tick species from which B. miyamotoi was amplified followed by the tick life stage/sex (NN = nymph; LL = larval pool), a four-letter alpha code indicating the avian host (AMRO = American Robin; HETH = Hermit Thrush; NOCA = Northern Cardinal), and a laboratory identification code. Additional sequences downloaded from GenBank comprise the reported genetic diversity of B. miyamotoi at the IGS locus and are labeled with spirochete origin.

Figure 4

Neighbor-joining phylogenetic tree based on 433 nucleotides of B. andersonii 16S-23S rRNA IGS haplotypes collected from Pitsfield Banding Station, 2004–2007. A sequence of B. burgdorferi obtained from GenBank was included as the outgroup. The percentages of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) are shown next to the branches when 80 or higher. Sequences are labeled with the name of the tick species from which B. andersonii was amplified followed by the tick life stage/sex (NN = nymph; LL = larval pool; AF = adult female) and a four-letter alpha code indicating the avian host (AMRO = American Robin; BHCO = Brown-headed Cowbird; BRTH = Brown Thrasher; CONW = Connecticut Warbler; ETTI = Eastern Tufted Titmouse; GRCA = Gray Catbird; HETH = Hermit Thrush; NOCA = Northern Cardinal; PUFI = Purple Finch; SOSP = Song Sparrow; SWTH = Swainson’s Thrush) or RABBIT indicating the eastern cottontail, a laboratory identification code, followed by the GenBank accession number. The single sequence derived from rabbit ear tissue is labeled with ‘EAR’. The asterisk and dagger denote samples from the same individual host.