Dataset Information

Paired real-time PCR assays for detection of Borrelia miyamotoi in North American Ixodes scapularis and Ixodes pacificus (Acari: Ixodidae).

ABSTRACT: Borrelia miyamotoi is an emerging, tick-borne human pathogen. In North America, it is primarily associated with Ixodes scapularis and Ixodes pacificus, two species known to bite humans. Here we describe the development and evaluation of a pair of real-time TaqMan PCR assays designed to detect B. miyamotoi in North American ticks. We sought to achieve sensitivity to B. miyamotoi strains associated with ticks throughout North America, the full genetic diversity of which is unknown, by targeting sequences that are largely conserved between B. miyamotoi strains from the eastern United States and genetically distinct B. miyamotoi strains from Japan. The two assays target different loci on the B. miyamotoi chromosome and can be run side by side under identical cycling conditions. One of the assays also includes a tick DNA target that can be used to verify the integrity of tick-derived samples. Using both recombinant plasmid controls and genomic DNA from North American and Japanese strains, we determined that both assays reliably detect as few as 5 copies of the B. miyamotoi genome. We verified that neither detects B. burgdorferi, B. lonestari or B. turicatae. This sensitive and specific pair of assays successfully detected B. miyamotoi in naturally-infected, colony-reared nymphs and in field-collected I. scapularis and I. pacificus from the Northeast and the Pacific Northwest respectively. These assays will be useful in screening field-collected Ixodes spp. from varied regions of North America to assess the risk of human exposure to this emerging pathogen.

SUBMITTER: Graham CB

PROVIDER: S-EPMC5572795 | biostudies-literature | 2016 Oct

REPOSITORIES: biostudies-literature

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Paired real-time PCR assays for detection of Borrelia miyamotoi in North American Ixodes scapularis and Ixodes pacificus (Acari: Ixodidae).

Graham Christine B CB Pilgard Mark A MA Maes Sarah E SE Hojgaard Andrias A Eisen Rebecca J RJ

Ticks and tick-borne diseases 20160718 6

Borrelia miyamotoi is an emerging, tick-borne human pathogen. In North America, it is primarily associated with Ixodes scapularis and Ixodes pacificus, two species known to bite humans. Here we describe the development and evaluation of a pair of real-time TaqMan PCR assays designed to detect B. miyamotoi in North American ticks. We sought to achieve sensitivity to B. miyamotoi strains associated with ticks throughout North America, the full genetic diversity of which is unknown, by targeting se ...[more]

PMID: 27475875

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Project description:Ixodes scapularis is a vector of several human pathogens in the United States, and there is geographical variation in the relative number of persons infected with these pathogens. Geographically isolated populations of I. scapularis have established or are in the process of establishing in southern Canada. Knowledge of the genetic variation within and among these populations may provide insight into their geographical origins in the United States and the potential risk of exposure of Canadians to the different pathogens carried by I. scapularis.Part of the mitochondrial (mt) 16S ribosomal (r) RNA gene was amplified by PCR from 582 ticks collected from southern Canada, and Minnesota and Rhode Island in the United States. Sequence variation was examined in relation to the predicted secondary structure of the gene. Genetic diversity among populations was also determined.DNA sequence analyses revealed 52 haplotypes. Most mutational alterations in DNA sequence occurred at unpaired sites or represented partial compensatory base pair changes that maintained the stability of the secondary structure. Significant genetic variation was detected within and among populations in different geographical regions. A greater proportion of the haplotypes of I. scapularis from the Canadian Prairie Provinces were found in the Midwest of the United States than in other regions, whereas more of the haplotypes of I. scapularis from the Canadian Central and Atlantic Provinces occurred in the Northeast of the United States. Nonetheless, 58% of I. scapularis were of a haplotype that occurs in the Midwest and Northeast of the United States; thus, their geographical origins could not be determined.There is considerable genetic variation in the mt 16S rRNA gene of I. scapularis. There is some evidence to support the hypothesis that some lineages of I. scapularis in the Atlantic and Central Provinces of Canada may be derived from colonizing individuals originating in the Northeast of the United States, whereas those in the Prairie Provinces may be derived from individuals originating in the Midwest of the United States. However, additional genetic markers are needed to test hypotheses concerning the geographical origins of I. scapularis in Canada.

Project description:Borrelia miyamotoi is a relapsing fever spirochete transmitted by ticks in the Ixodes ricinus complex. In the eastern United States, B. miyamotoi is transmitted by I. scapularis, which also vectors several other pathogens including B. burgdorferi sensu stricto. In contrast to Lyme borreliae, B. miyamotoi can be transmitted vertically from infected female ticks to their progeny. Therefore, in addition to nymphs and adults, larvae can vector B. miyamotoi to wildlife and human hosts. Two widely varying filial infection prevalence (FIP) estimates - 6% and 73% - have been reported previously from two vertically infected larval clutches; to our knowledge, no other estimates of FIP or transovarial transmission (TOT) rates for B. miyamotoi have been described in the literature. Thus, we investigated TOT and FIP of larval clutches derived from engorged females collected from hunter-harvested white-tailed deer in 2015 (n = 664) and 2016 (n = 599) from Maine, New Hampshire, Tennessee, and Wisconsin. After engorged females oviposited in the lab, they (n = 492) were tested for B. miyamotoi infection by PCR. Subsequently, from each clutch produced by an infected female, larval pools, as well as 100 individual eggs or larvae, were tested. The TOT rate of the 11 infected females was 90.9% (95% CI; 57.1-99.5%) and the mean FIP of the resulting larval clutches was 84.4% (95% CI; 68.1-100%). Even though the overall observed vertical transmission rate (the product of TOT and FIP; 76.7%, 95% CI; 44.6-93.3%) was high, additional horizontal transmission may be required for enzootic maintenance of B. miyamotoi based on the results of a previously published deterministic model. Further investigation of TOT and FIP variability and the underlying mechanisms, both in nature and the laboratory, will be needed to resolve this question. Meanwhile, studies quantifying the acarological risk of Borrelia miyamotoi disease need to consider not only nymphs and adults, but larval I. scapularis as well.

Dataset Information

Paired real-time PCR assays for detection of Borrelia miyamotoi in North American Ixodes scapularis and Ixodes pacificus (Acari: Ixodidae).

Publications

Paired real-time PCR assays for detection of Borrelia miyamotoi in North American Ixodes scapularis and Ixodes pacificus (Acari: Ixodidae).

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