A comparative analysis of molecular markers for the detection and identification

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J Med Microbiol 59 (2010), 309-314; DOI: 10.1099/jmm.0.013508-0

A comparative analysis of molecular markers for the detection and

identification of Borrelia spirochaetes in Ixodes ricinus

Beata Wodecka, Agata Leoska and Bogumia Skotarczak

Department of Genetics, University of Szczecin, 71-065 Szczecin, Poland

Correspondence

Agata Leoska

_agataleonska_ (agataleonska)

Received June 10, 2009

Accepted December 9, 2009

 

Borrelia burgdorferi sensu lato, carried by Ixodes ticks, is one of the

most significant human pathogens, causing Lyme disease. As there is no

standardized PCR method for detection and identification of spirochaete DNA, we

carried out a comparative analysis using a set of complementary primers

for three regions in the genomic DNA of these bacteria (genes fla and rrs and

the non-coding rrs–rrlA region). DNA extracted from 579 Ixodes ricinus

ticks was subjected to nested PCR. DNA of the examined spirochaetes was detected

in 43 (7.4 %) lysates when the fla gene was used as a molecular marker, in

7 (1.2 %) lysates when using primers complementary to the rrs gene, and in

12 (2.1 %) lysates using primers complementary to the non-coding rrs–rrlA

sequence. RFLP analysis based on the fla gene helped identify species from the

B. burgdorferi sensu lato complex (B. burgdorferi sensu stricto, Borrelia

afzelii, Borrelia garinii, Borrelia valaisiana), detect co-infections, and

also identify Borrelia miyamotoi. Therefore, the fla gene is the most

sensitive and specific molecular marker for the detection and identification of

Borrelia spirochaetes in I. ricinus.

The GenBank/EMBL/DDBJ accession numbers for the fla gene sequence of B.

burgdorferi sensu stricto G112-04, B. afzelii D131-07, B. garinii G23-07, B.

valaisiana G43-0507 and B. miyamotoi D110-07 are FJ518808, FJ518805,

FJ518806, FJ518807 and FJ518804, respectively.