Sequence and secondary structure of the mitochondrial 16S ribosomal RNA gene of Ixodes scapularis.

The complete DNA sequences and secondary structure of the mitochondrial (mt) 16S ribosomal (r) RNA gene were determined for six Ixodes scapularis adults. There were 44 variable nucleotide positions in the 1252 bp sequence alignment. Most (95%) nucleotide alterations did not affect the integrity of the secondary structure of the gene because they either occurred at unpaired positions or represented compensatory changes that maintained the base pairing in helices. A large proportion (75%) of the intraspecific variation in DNA sequence occurred within Domains I, II and VI of the 16S gene. Therefore, several regions within this gene may be highly informative for studies of the population genetics and phylogeography of I. scapularis, a major vector of pathogens of humans and domestic animals in North America.

Sequence variability in the mitochondrial 12S rRNA and tRNA Val genes of Ixodes scapularis (Acari: Ixodidae) individuals shown previously to be genetically invariant.

The DNA sequences of the mitochondrial (mt) 12S rRNA and tRNA(Val) genes were characterized for 82 blacklegged ticks (Ixodes scapularis) that were genetically identical for Domains IV and V of the mt 16S rRNA gene. Thirty-one haplotypes, differed in sequence by 1-9 bp, were detected among the 82 ticks. Most nucleotide alterations in DNA sequence did not affect the stability of the secondary structures of the RNAs. The magnitude of the DNA sequence variation in the mt 12S rRNA and tRNA(Val) genes among blacklegged ticks suggests that this region of the mitochondrial genome has potential as a genetic marker for examining the population genetics and phylogeography of I. scapularis.

Two Anaplasma phagocytophilum strains in Ixodes scapularis ticks, Canada.

We developed PCR-based assays to distinguish a human pathogenic strain of Anaplasma phagocytophilum, Ap-ha, from Ap-variant 1, a strain not associated with human infection. The assays were validated on A. phagocytophilum-infected black-legged ticks (Ixodes scapularis) collected in Canada. The relative prevalence of these 2 strains in I. scapularis ticks differed among geographic regions.

Granulocytic anaplasmosis in a horse from Saskatchewan.

This report describes a case of equine granulocytic anaplasmosis in a horse from Saskatchewan. Morulae were visualized within blood neutrophils, and the diagnosis was confirmed by polymerase chain reaction (PCR). The organism was identified as the human pathogenic strain of Anaplasma phagocytophilum by PCR and DNA sequencing of 3 independent genes.

RésuméAnaplasmose granulocytaire chez un cheval de la Saskatchewan. Ce rapport décrit un cas d'anaplasmose granulocytaire chez un cheval de la Saskatchewan. Des morulas ont été visualisées dans les neutrophiles sanguins et le diagnostic a été confirmé par une réaction d'amplificaton en chaîne par la polymérase (PCR). L'organisme a été identifié comme la souche pathogène humaine d'Anaplasma phagocytophilum par PCR et séquençage d'ADN de 3 gènes indépendants.(Traduit par Isabelle Vallières).

An assessment of genetic differences among ixodid ticks in a locus within the nuclear large subunit ribosomal RNA gene.

We examined the usefulness of the D3 domain and flanking core regions (=D3(+)) of the nuclear large subunit (LSU) ribosomal DNA as a genetic marker for species-level identification and the inference of evolutionary relationships of ixodid ticks. Genetic variation was also examined in relation to the secondary structure of the LSU rDNA. The results revealed a lack of sequence difference in the D3(+) among species of Dermacentor and among some species of Ixodes, demonstrating that this gene region is not suitable as a species marker for all species of ixodid ticks. Of the 45 variable nucleotide positions in the sequence alignment of the D3(+), 23 did not alter the secondary structure of the LSU rDNA, because they occurred in unpaired positions, whereas 16 represented partial or full compensatory changes which maintained the secondary structure. Six deletions in the D3(+) sequence of all Ixodes species examined resulted in a shorter d4_1 helix compared with that of other tick species. The results of the phylogenetic analyses also showed that the D3(+) is of limited value in resolving evolutionary relationships among ixodid ticks. In addition, we also demonstrated that the D3(+) of ascomycete fungi could also be amplified along with, or instead of, the D3(+) of some tick species, depending upon the primers used in PCR. Nonetheless, the D3(+) of the fungal contaminants are readily distinguished from the D3(+) of ixodid ticks because of a shorter length and the absence of helix d4_1 in the secondary structure of the LSU rDNA.

Genetic variation in the mitochondrial 16S ribosomal RNA gene of Ixodes scapularis (Acari: Ixodidae).

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

Genetic diversity in Ixodes scapularis (Acari: Ixodidae) from six established populations in Canada.

Although Ixodes scapularis is the most important vector of Borrelia burgdorferi sensu stricto, the causative agent of Lyme disease in North America, little is known of the genetic diversity in this tick species within the recently established populations in Canada. In the present study, 153 I. scapularis adults collected from southern Canada were compared genetically using single-strand conformation polymorphism (SSCP) analyses in combination with DNA sequencing of the mitochondrial 16S rRNA gene. Nineteen haplotypes were detected, 8 of which have not been reported in the U.S.A. One 'new' haplotype was only detected at Lunenburg, Nova Scotia, and comprised 38% of the ticks examined for that population. The population in the southeast corner of Manitoba contained 3 'new' haplotypes. Although the most common haplotype (Is-1) was present in all 6 populations of I. scapularis in Canada, there were significant differences in the genetic structure among population. This suggests different geographical origins for the tick populations in Canada, which may be related to the transportation of larval and nymphal ticks by migratory passerines using different flyways. Determination of the origins of the endemic populations of I. scapularis in Canada, as well as those predicted to establish in the near future, has important implications with respect to understanding the risk of exposure to pathogenic bacteria infecting these ticks.

Genetic variation in the mitochondrial 16S rRNA gene of the American dog tick, Dermacentor variabilis (Acari: Ixodidae).

Genetic variation in the mitochondrial (mt) 16S ribosomal RNA (rRNA) gene was examined for the American dog tick, Dermacentor variabilis (Say, 1821). Nine different haplotypes were detected among 369 adult D. variabilis collected from four localities in Canada. There were eight variable nucleotide positions in the 404 bp sequence alignment. Individuals of haplotype 1 occurred at frequency of >75% at all localities. Five haplotypes were detected at only one of the four localities. High haplotype diversity and low nucleotide diversity, combined with significantly negative Fs values for ticks at three localities, suggest a recent population expansion. Genetic differences were found between populations at different localities, but a Mantel regression analysis revealed no association between genetic differences and geographical distances. There was also no association between tick haplotype and the prevalence of the bacterium, Rickettsia montanensis Weiss and Moulder, 1984, in D. variabilis among localities or on opposite sides of Blackstrap Lake (Saskatchewan). The 16S rDNA haplotypes from Canadian populations of D. variabilis formed a clade with those from the eastern and central U.S.A., to the exclusion of D. variabilis from geographically isolated populations in the western U.S.A. Although sample sizes for D. variabilis in the eastern U.S.A. are small, there may be genetic divergence between populations in Canada and those in the eastern U.S.A., which may have implications for studies on the pathogenic agents transmitted by D. variabilis to its hosts.