Original article
Detection of a novel Rickettsia (Alphaproteobacteria: Rickettsiales) in rotund ticks (Ixodes kingi) from Saskatchewan, Canada

https://doi.org/10.1016/j.ttbdis.2012.11.013Get rights and content

Abstract

A novel Rickettsia was detected in the rotund tick, Ixodes kingi Bishopp, 1911, based on comparative DNA sequence analyses of 4 genes; the rickettsial-specific 17-kDa antigen gene, citrate synthase gene (gltA), the outer surface membrane protein A gene (ompA), and the 16S rRNA gene. The rickettsiae in I. kingi differed in nucleotide sequence from those of other Rickettsia species by 5.8–18.3% for the 17-kDa gene, 0.9–13.9% for gltA, 5.5–22.8% for ompA, and 0.9–1.6% for the 16S rRNA gene. Phylogenetic analyses of the sequence data revealed that this putative new species of Rickettsia, provisionally named Candidatus Rickettsia kingi, does not belong to the spotted fever group or typhus group of rickettsiae, but represents a sister taxon to R. canadensis and Candidatus Rickettsia tarasevichiae. This novel Rickettsia was found in 60 of the 87 (69%) ticks examined, which included all feeding life cycle stages of I. kingi. Although adult I. kingi occasionally parasitize dogs and humans, it remains to be determined if this Rickettsia is pathogenic to these host species.

Introduction

Rickettsiae are obligate intracellular bacteria that are transmitted to vertebrates by arthropod vectors that include ticks, fleas, lice, and mites (Fournier and Raoult, 2009, Merhej and Raoult, 2011). These Alphaproteobacteria are the causative agents of disease (e.g., spotted fever and typhus) in many parts of the world (Fournier and Raoult, 2009, Merhej and Raoult, 2011). There are at least 30 recognized species of Rickettsia, 19 of which are considered human pathogens (Merhej and Raoult, 2011). A number of other putative species of Rickettsia have also been proposed based on sequence differences in 2 or more genes (e.g., Almeida et al., 2011, Izzard et al., 2009, Pacheco et al., 2011, Phan et al., 2011, Shpynov et al., 2003). Historically, the genus Rickettsia has been divided into the spotted fever group (SFG), the typhus group (TG), R. canadensis, and R. bellii (Fournier and Raoult, 2009). The SFG contains the majority of species within the genus, while R. typhi and R. prowazekii are the members of the TG (Fournier and Raoult, 2009). The TG is associated primarily with lice and fleas, whereas R. canadensis, R. bellii, and the SFG (except for R. akari and R. felis) use ixodid ticks as vectors (Fournier and Raoult, 2009, Merhej and Raoult, 2011).

In North America, at least 6 species of Ixodes (i.e., I. scapularis, I. pacificus, I. cookei, I. dentatus, I. brunneus, and I. texanus), all of which are known to parasitize rodents (Allan, 2001, Bishopp and Trembley, 1945, Kolonin, 2007), have been shown to contain SFG rickettsiae (Allan, 2001, Anderson et al., 1986, Billings et al., 1998, Clifford et al., 1969, Magnarelli et al., 1985, Phan et al., 2011). The rotund tick, Ixodes kingi, is also a common parasite of rodents (i.e., murids, heteromyids, geomyids, and sciurids), as well as other vertebrates, in western North America (Allan, 2001, Bishopp and Trembley, 1945, Gregson, 1971, Salkeld et al., 2006). Although rotund ticks are known to be vectors of several pathogens, including Coxiella burnetii, the causative agent of Q fever, and Francisella tularensis, the causative agent of tularemia (Sidwell et al., 1964, Thorpe et al., 1965), there are no published reports of rickettsiae in I. kingi. In this paper, we report the discovery of a new species of Rickettsia in all feeding life cycle stages of I. kingi from a locality in central Saskatchewan, Canada.

Section snippets

Materials and methods

For this study, a total of 87 I. kingi (3 females, 1 male, 2 nymphs, and 81 larvae) were collected from northern pocket gophers (Thomomys talpoides) trapped near Clavet, Saskatchewan (Anstead and Chilton, 2011). Total genomic (g) DNA was extracted from each tick as described by Dergousoff and Chilton (2007). The presence of rickettsiae in ticks was determined by nested (n)-PCR targeting a 434-bp fragment of the rickettsial-specific 17-kDa antigen gene using primers 17k-5 and 17k-3 (first phase)

Results

A single band of the expected size (∼450 bp) for the partial rickettsial 17-kDa gene was detected on agarose gels for amplicons derived from the gDNA of 60 of the 87 (69%) ticks. These 60 PCR-positive samples represented all feeding life cycle stages of I. kingi (i.e., 55 larvae, 1 nymph, and 4 adults). No bands were detected on agarose gels for the negative control (i.e., no gDNA) samples. The banding patterns of all 60 PCR-positive samples on SSCP gels were identical to one another, but

Discussion

This study, as far as we are aware, represents the first published record of the detection of Rickettsia within I. kingi. Furthermore, this bacterium, present within all feeding life cycle stages of I. kingi, represents a novel species of Rickettsia because it differs in DNA sequence at 4 gene loci (i.e., 17-kDa gene, gltA, ompA, and 16S rRNA gene) when compared to the sequences of all recognized and putative species within the genus. The magnitude of sequence differences of this novel

Acknowledgments

This work was approved by the University of Saskatchewan's Animal Research Ethics Board, and adhered to the Canadian Council on Animal Care guidelines for humane animal use. Financial support for this work was provided (to NBC) from the Natural Sciences and Engineering Research Council of Canada and the Canadian Foundation for Innovation. A Margaret McKay scholarship and a University of Saskatchewan Graduate Scholarship provided financial support to CAA. We are grateful to Dr. C Todd for his

References (32)

  • A.P. Almeida et al.

    A novel Rickettsia infecting Amblyomma dubitatum ticks in Brazil

    Ticks Tick-Borne Dis.

    (2011)
  • S.J. Dergousoff et al.

    Differentiation of three species of ixodid tick, Dermacentor andersoni, D. variabilis and D. albipictus, by PCR-based approaches using markers in ribosomal DNA

    Mol. Cell. Probes

    (2007)
  • S.A. Allan

    Ticks (Class Arachnida: Order Acarina)

  • J.F. Anderson et al.

    Rickettsia rickettsii and Rickettsia montana from ixodid ticks in Connecticut

    Am. J. Trop. Med. Hyg.

    (1986)
  • C.A. Anstead et al.

    Ticks feeding on northern pocket gophers (Thomomys talpoides) in central Saskatchewan and the unexpected detection of Ixodes scapularis larvae

    J. Vector Ecol.

    (2011)
  • A.N. Billings et al.

    Molecular characterization of a novel Rickettsia species from Ixodes scapularis in Texas

    Emerg. Infect. Dis.

    (1998)
  • F.C. Bishopp et al.

    Distribution and hosts of certain North American ticks

    J. Parasitol.

    (1945)
  • F.M. Bozeman et al.

    Serologic evidence of Rickettsia canada infection of man

    J. Infect. Dis.

    (1970)
  • C.M. Clifford et al.

    Tests on ticks from wild birds collected in the eastern United States for rickettsiae and viruses

    Am. J. Trop. Med. Hyg.

    (1969)
  • S.J. Dergousoff et al.

    Association of different genetic types of Francisella-like organisms with the Rocky Mountain wood tick (Dermacentor andersoni) and the American dog tick (Dermacentor variabilis) in localities near their northern distributional limits

    Appl. Environ. Microbiol.

    (2012)
  • S.J. Dergousoff et al.

    Prevalence of Rickettsia species in Canadian populations of Dermacentor andersoni and D. variabilis

    Appl. Environ. Microbiol.

    (2009)
  • M.E. Eremeeva et al.

    Detection and identification of bacterial agents in Ixodes persulcatus Schulze ticks from the north western region of Russia

    Vector-Borne Zoonotic Dis.

    (2007)
  • P.-E. Fournier et al.

    Current knowledge on phylogeny and taxonomy of Rickettsia spp.

    Ann. N. Y. Acad. Sci.

    (2009)
  • R.B. Gasser et al.

    Single-strand conformation polymorphism (SSCP) for the analysis of genetic variation

    Nat. Protoc.

    (2006)
  • J.D. Gregson

    Studies on two populations of Ixodes kingi Bishopp (Ixodidae)

    Can. J. Zool.

    (1971)
  • S.R. Heise et al.

    Bacterial diversity in Amblyomma americanum (Acari: Ixodidae) with a focus on members of the genus Rickettsia

    J. Med. Entomol.

    (2010)
  • Cited by (24)

    • Multilocus genetic analysis indicates taxonomic status of “Candidatus Rickettsia mendelii” as a separate basal group

      2023, Ticks and Tick-borne Diseases
      Citation Excerpt :

      The R. canadensis group includes several species: R. canadensis, “Candidatus Rickettsia tarasevichiae”, Rickettsia monteiroi, “Candidatus Rickettsia angustus”, “Candidatus Rickettsia kingi”, and Rickettsia endosymbiont of Amblyomma tonelliae. The Rickettsia bellii group comprises several R. bellii strains (Anstead et al., 2013a, b; Pacheco et al., 2011; Tarragona et al., 2015). Rickettsia spp. belonging to the R. canadensis and R. bellii groups and the most part of SFG rickettsiae (SFGR) are transmitted by ixodid ticks.

    • Emerging spotted fever group rickettsiae in ticks, northwestern China

      2016, Ticks and Tick-borne Diseases
      Citation Excerpt :

      Phylogenetic trees were constructed using the maximum likelihood (ML) and neighbor joining (NJ) algorithms with MEGA 6.0 (Tamura et al., 2013). Bootstrap analyses (500 replicates for ML analyses and 1000 replicates for the NJ analyses) were conducted to determine the relative support for clades in the consensus trees (Pattengale et al., 2010; Anstead and Chilton, 2013a,b). A total of 117 Rh.

    • Molecular detection of spotted fever group rickettsiae in ticks removed from humans in Turkey

      2016, Ticks and Tick-borne Diseases
      Citation Excerpt :

      In the parallel, numerous ecological and epidemiological studies about tick-borne rickettsial infections have been conducted throughout the world (Parola et al., 2005, 2013; Oteo and Portillo, 2012). During this period, many new pathogenic or nonpathogenic rickettsiae have been also described or isolated from ticks (Beati et al., 1997; Fournier et al., 2006; Palomar et al., 2012; Anstead and Chilton, 2013; Kurtti et al., 2015). Rickettsial infections have been very poorly studied in Turkey.

    • Mutation scanning-based identification of larval and nymphal ticks (Acari: Ixodidae) from Richardson's ground squirrels (Spermophilus richardsonii)

      2014, Molecular and Cellular Probes
      Citation Excerpt :

      In the present study, PCR-based single-strand conformation polymorphism (SSCP) analyses and DNA sequencing of the mt 16S rRNA gene were used to identify larval and nymphal ticks feeding on S. richardsonii in southwestern Saskatchewan (Canada). SSCP is a mutation scanning technique that can effectively display genetic variation within or among DNA sequences of up to 450 bp in size [21]; its combined use with DNA sequencing, employing a suitable genetic marker, has been shown to be a useful approach for the specific identification of a variety of organisms (e.g., [7,9,22–26]). A total of 116 ticks (24 adults, 48 nymphs, 44 larvae) were removed from 18 (44%) of 41 S. richardsonii collected during the summer months (June and July) of 2009 and 2010 from a cattle farm situated on the outskirts of the village of Beechy, Saskatchewan (50.8833N, 107.3833W).

    View all citing articles on Scopus
    View full text