Bartonella Species Isolated from Rodents, Greece

摘要

To the Editor: Domestic cats andhuman body lice have been identifiedas the vectors of Bartonella henselaeand B. quintana, respectively, the pri-mary sources of Bartonella-associat-ed human diseases (1). Bartonellaspecies are zoonotic agents that havebeen isolated from a wide range ofmammals in the United States (2) andEurope (3) and have been associatedwith human diseases (4–5). This study investigated the poten-tial for infection from Bartonellaspecies in rodents in northern Greece.The small mammals tested were col-lected with live traps (6). Two siteswere surveyed; the first wasNevrokopi, a small town in theRhodope Mountains near the Greek-Bulgarian border, and the second siteincluded Pramanta, a small village inthe Pindos Mountains, and Matsuki, asmall village in northwestern Greece.At Nevrokopi, 57 small mammalswere captured during 887 trap nightsfor a success rate of 6.4%. AtPramanta and Matsuki, 13 smallmammals were captured during 400trap nights for a success rate of 3.3%.The 70 captured mammals comprisedseven species of rodents. Apodemusflavicollis was the most commonlycaptured species (87%). Blood sam-ples from each of the trapped mam-mals were frozen in liquid nitrogen inthe field and subsequently stored at–70oC before bacteria isolation.Bacteria isolation was performed aspreviously described (7). One hun-dred microliters of whole mammalianblood was cultured on heart infusionagar containing 5% rabbit blood(Becton Dickinson, Franklin Lakes,NJ) and incubated in 5% CO2at 35°Cfor a minimum of 4 weeks. DNA ofthe putative Bartonella cultures wasextracted by using QIAamp TissueKit (Qiagen GmbH, Hilden,Germany). Polymerase chain reaction(PCR) was performed by using twooligonucleotides specific for the cit-rate synthase (gltA) gene of B. hense-lae Houston 1, primers BhCS 781.pand BhCS 1137.n. Negative and posi-tive controls (double-distilled H2Oand DNA from cultures of B. hense-lae) were used in each PCR run.Products of the correct size were puri-fied (QIAquick PCR Purification kit,Qiagen GmbH) and sequenced withthe same primers, BhCS 781.p andBhCS1137.n., in both directions, withthe Cy5/Cy5.5 Dye Primer CycleSequencing kit on a Long-ReadTower sequencer (Visible GeneticsInc., Toronto, Canada). Three hundredthirty-eight base-pair sequences of thegltA gene were obtained and com-pared with sequences of other knownBartonella species in GenBank byusing the nucleotide BLAST program(National Center for BiotechnologyInformation; Available from:www.ncbi.nlm.nih.gov/BLAST/).Isolates identified as Bartonellaspecies were obtained from 21 of the70 blood cultures. All were isolatedfrom A. flavicollis, and one was isolat-ed from Dryomys nitedula. In addi-tion, all were isolated from the firstsite (Nevrokopi village), and one wasisolated from the second site(Pramanta village).