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Bacteriology

Limited Diversity among Human Isolates of Bartonella henselae

B. Dillon, J. Valenzuela, R. Don, D. Blanckenberg, D. I. Wigney, R. Malik, A. J. Morris, J. M. Robson, J. Iredell
B. Dillon
1Centre for Infectious Diseases and Microbiology, Westmead Hospital, University of Sydney, New South Wales 2145
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J. Valenzuela
1Centre for Infectious Diseases and Microbiology, Westmead Hospital, University of Sydney, New South Wales 2145
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R. Don
1Centre for Infectious Diseases and Microbiology, Westmead Hospital, University of Sydney, New South Wales 2145
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D. Blanckenberg
1Centre for Infectious Diseases and Microbiology, Westmead Hospital, University of Sydney, New South Wales 2145
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D. I. Wigney
2Faculty of Veterinary Science, University of Sydney, New South Wales 2006
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R. Malik
2Faculty of Veterinary Science, University of Sydney, New South Wales 2006
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A. J. Morris
3Department of Microbiology, Green Lane Hospital, Auckland 1003, New Zealand
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J. M. Robson
4Sullivan and Nicolaides Pathology, Taringa, Queensland 4068, Australia
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J. Iredell
1Centre for Infectious Diseases and Microbiology, Westmead Hospital, University of Sydney, New South Wales 2145
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  • For correspondence: joni@icpmr.wsahs.nsw.gov.au
DOI: 10.1128/JCM.40.12.4691-4699.2002
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  • FIG. 1.
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    FIG. 1.

    Representative gel electrophoresis of 16S rDNA type-specific PCR. Lanes 1 to 9, BH1 type-specific PCR; lanes 10 to 18, BH2 type-specific PCR. Lanes 1 and 10, JR1; lanes 2 and 11, ATCC 49793; lanes 3 and 12, ATCC 49882 (Houston-1); lanes 4 and 13, URLLY-8 (Marseille); lanes 5 and 14, R1073; lanes 6 and 15, NU4423; lanes 7 and 16, HC35; lanes 8 and 17, NU4428; lanes 9 and 18, negative (water). M, 0.07- to 12.2-kbp marker (Roche Diagnostics, Mannheim, Germany).

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    FIG. 2.

    (A) Representative HaeIII restriction fragment length polymorphism patterns of PCR-amplified 16S-23S ITS region. Lane 1, JR2; lane 2, JR3; lane 3, URLLY-8 (Marseille); lane 4, ATCC 49882 (Houston-1); lane 5, ATCC 49793; lane M, 0.07- to 12.2-kbp marker (Roche) (sizes are indicated on the right-hand margin); lane 6, HC63; lane 7, NU4428; lane 8, NU4714; lane 9, RMC1; lane 10, negative control (water). (B) 16S-23S rRNA ITS sequence of B. henselae Houston-1 (GenBank accession no. L35101 ). The tRNA, 16S, and 23S rRNA genes are underlined. The repeat region is boxed, and arrows denote base substitutions or insertions which vary from the Houston-1 sequence, as follows: 1, G-to-A substitution in HC35, HC54, HC60, HC62, HC69, NU4681, NU4714, JR2, BH2, R987, and R1073; 2, G-to-A substitution in HC54, HC69, NU4681, JR2, and BH2; 3, T-to-G substitution in HC35 and HC69; 4, C-to-T substitution in HC69; 5, C-to-T substitution in HC35, HC60, HC62, NU4428, NU4681, NU4714, RMC1, RMC8, RMC12, R987, and R1073; 6, A-to-G substitution in HC35, HC54, HC60, HC62, HC69, NU4428, NU4714, RMC1, RMC3, RMC8, RMC10, RMC12, R1073, R987, and JR2; 7, G-to-A substitution in HC60, HC62, NU4428, NU4714, RMC1, RMC8, RMC12, R987, and R1073. (C) 16S-23S ITS repeat region. There are two ATTGCTTCTAAAAAG repeats in HC54, HC69, RMC3, and JR2; three repeats in RMC10; and one repeat in BH2. The boxed region illustrates the previously described repeat sequence in B. henselae SA-2 (14).

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    FIG. 3.

    AP-PCR representative gel. Lane 1, ATCC 49882 (Houston-1); lane 2, JR2; lane 3, RMC1; lane 4, BH2; lane 5, RMC8; lane 6, URLLY-8 (Marseille); lane 7, HC63; lane 8, HC77; lane 9, BH4; lane 10, NU4428; M, pGEM marker (Promega; molecular size in base pairs is given along the right border).

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    FIG. 4.

    ERIC-PCR. (A) Representative gel electrophoresis of ERIC-PCR. Lanes 1 to 3, pattern E4H (strains ATCC 49882 [Houston-1] and NU4714); lanes 4 and 5, pattern E4M (strains BH4 and Marseille); lanes 6 and 7, pattern E2 (strains HC61 and NU4428); lanes 8 and 9, patterns E3a and -b, respectively (strains HC35 and HC60); lanes 10 and 11, pattern E1 (strains HC62 and RMC1); lanes 12 and 13, pattern E5 (strains RMC3 and RMC7); lanes 14 and 15, pattern E6 (strain RMC2 duplicate); lanes 16 and 17, pattern E7 (strains RMC11 and RMC4); lanes M1 and M2, molecular size markers. (B) Dendrogram. B. quintana and B. vinsonii patterns are quite distinct from all others (not shown in panel A) and are included in a merged reconstructed dendrogram. The dendrogram reflects the similarity of distribution of ERIC priming sites and is derived from gels (including that shown in panel A), all of which included B. henselae and the two molecular size markers and were analyzed identically.

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    FIG. 5.

    EagI-HhaI IRS-PCR. Lane 1, negative control; lane 2, NU4713; lane 3, HC61; lane 4, BH2; lane 5, R1073; lane 6, Marseille; lane 7, ATCC 49882 (Houston-1); lane 8, NU4714; lane 9, ATCC 49793; lane 10, HC35; lane 11, HC60; lane M, pGEM marker (Promega) (molecular sizes are given along the right border).

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  • TABLE 1.

    B. henselae isolates studieda

    Isolate(s)Source16S typegltA typeAP typeERIC typeIRS type
    Feline isolates (n = 31)
        NU4695BloodIHHE4H1
        RMC10†BloodIHHE4H1
        HC54BloodIHHE4H2
        HC69BloodIHME4H2
        HC62†BloodIHHE12
        RMC3*BloodIHHE52
        NU4428†BloodIHME22
        HC71BloodIMHE22
        NU4714*†BloodIIHHE4H1
        HC35*, HC55BloodIIHHE3a1
        RMC8*BloodIIHHaE3a2
        NU4681*BloodIIHHE3a1
        HC60†BloodIIHHE3b1
        RMC1*BloodIIHHE12
        RMC12BloodIIHME3a1
        NU4423†BloodIIMHE4H2
        RMC11BloodIIMME71
        RMC4BloodIIMME72
        HC41, HC48†, HC61, HC63†, HC77BloodIIMME22
        RMC2†BloodIIMME62
        RMC5BloodIIMME12
        RMC6†, RMC9†BloodIIMME52
        RMC7, Berlin-2BloodIIMME52
        NU4713BloodIIMHE22
    Human isolates (n = 28)
        BH2CSDIHHE4H1
        ATCC 49793/NZRm3492 (s4)BAIHHE4H1
        ATCC 49882 (Houston-1)†, Berlin-1BAIHHE4H2
        BH3(s4), BH5s2*, R987, JR1, JR2, JR3, JR5, JR6, JR7, JR8, JR9, JR12, JR13, JR14, JR15, JR17, JR18, JR19, JR20, JR22, JR23CSDIHHE4H2
        R1073†CSDIIHHE4H2
        BH4CSDIIMME4M2
        URLLY-8 (Marseille)†BAIIMME4M2
    • ↵ a Symbols: *, Isolate embeds into solid agar and displays ratchet motility at this passage number; †, 16S type was determined by direct sequencing. s2, second passage after initial isolation; (s4), third to fifth passage after initial isolation. If not specified, RMC-prefix isolates (this work) were studied at passage numbers 2 to 3; HC-prefix isolates were a gift of Tom Gottlieb (8) and were studied at passage numbers 4 to 6; NU-prefix isolates (15) were studied at passage numbers 3 to 5; R- and JR-prefix strains (this work and Fournier et al. [12a]) were all studied at passage numbers 4 to 7. Other isolates were isolated in our laboratory or obtained from the following: Mardjan Arvand, Hygiene-Institute, Heidelberg, Germany (2) (Berlin-1 and -2); R. Bunter, Goulburn Valley Base Hospital, Goulburn, NSW, Australia (BH2); the New Zealand quality control strain NZRm3492 is thought to be a lab-passaged derivative of the widely used strain ATCC 49793, and they are inseparable by all typing methods; Jock Harkness, Royal North Shore Hospital, Sydney, Australia [BH3(s4)]; T.H.E. Pathology, Fairfield Heights, NSW, Sydney, Australia [BH5(S2)]; and D. Raoult, Unite des Rickettsies, Marseille, France (9) (URLLY-8 [Marseille]). Strains with the ATCC prefix were obtained from the American Type Culture Collection, Manassas, Va. 16S-23S regions were sequenced in all feline strains, in all Sydney and imported human strains, and in JR2.

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Limited Diversity among Human Isolates of Bartonella henselae
B. Dillon, J. Valenzuela, R. Don, D. Blanckenberg, D. I. Wigney, R. Malik, A. J. Morris, J. M. Robson, J. Iredell
Journal of Clinical Microbiology Dec 2002, 40 (12) 4691-4699; DOI: 10.1128/JCM.40.12.4691-4699.2002

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Limited Diversity among Human Isolates of Bartonella henselae
B. Dillon, J. Valenzuela, R. Don, D. Blanckenberg, D. I. Wigney, R. Malik, A. J. Morris, J. M. Robson, J. Iredell
Journal of Clinical Microbiology Dec 2002, 40 (12) 4691-4699; DOI: 10.1128/JCM.40.12.4691-4699.2002
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KEYWORDS

Bartonella henselae
Cat Diseases
Cat-Scratch Disease
Genetic Variation

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