Emergence of Related Nontoxigenic Corynebacterium Biotype mitis Strains in Western Europe.

We report on 17 isolates of Corynebacterium diphtheriae biotype mitis with [ILLEGIBLE TEXT] from Switzerland, Germany, and France. Isolates came from skin and [ILLEGIBLE TEXT] of injecting drug users, homeless persons, prisoners, and elderly orthopedic [ILLEGIBLE TEXT] prostheses or primary joint infections. Such isolates had only been observed in

Nontoxigenic Corynebacterium diphtheriae strains were recovered from [ILLEGIBLE TEXT] throat swabs from immunized British military personnel in Germany from 1993 [ILLEGIBLE TEXT] nontoxigenic C. diphtheriae biotype mitis isolates had been described in skin, [ILLEGIBLE TEXT] of Swiss injecting drug users; 32 of the isolates belonged to the same clone (2,3). demonstrates that this clone and closely related clones occurred between 1990 an European countries, and only sometimes in persons with poor hygiene.

Five C. diphtheriae isolates came from two laboratories in Zurich and Bern, Swit laboratories in Hamburg, Germany; and 1 from Paris, France (Table). They were C. diphtheriae biotype mitis (4); that is, they were nonlipophilic, were nitrate red did not ferment glycogen. By polymerase chain reaction techniques (5), the [ILLEGIBLE TEXT] not detected in any isolate. For some isolates, the Elek test was also performed; [ILLEGIBLE TEXT] negative. For ribotyping, DNA was isolated, digested with either EcoRI or PvuII, blotted, and probed for rDNA as described elsewhere (2,3). Disk susceptibility [ILLEGIBLE TEXT] and MIC determinations were done according to National Committee for Clinical methods (6). All other bacteria isolated were also identified and serotyped in [ILLEGIBLE TEXT]

Table. Nontoxigenic Corynebacterium diphtheriae isolates

Isolate   Date          Place of
no.       isolated(a)   isolation(b)

2012      1990          Paris

2410      2/1995        Hamburg

2689      3/1995        Hamburg

1682      9/1995        Hamburg

1935      3/1996        Hamburg

1836      3/1996        Hamburg

2661      3/1996        Hamburg

2658      4/1996        Hamburg

2674      5/1996        Hamburg

2670      7/1996        Hamburg

2667      7/1996        Hamburg

2413      11/1996       Hamburg

2446      12/1996       Zurich

2464      1/1997        Zurich

2473      1/1997        Zurich

2475      1/1997        Zurich

 480      5/1997        Bern

               Ribotyping
               pattern(c)

                          Patients
Isolate                   sex,
no.       EcoRI   PvuII   age (yr)

2012        A       C      m, 6

2410        A       B      m, 30

2689        A       A      f, 82

1682        A       B      m, 45

1935        A       B      m, 58

1836        A       B      m, 45

2661        A       A      m, 35

2658        A       A      f, 76

2674        A       A      f, 62

2670        A       A      f, 81

2667        A       A      m, 23

2413        A       B      m, 36

2446        A       A      f, 38

2464        A       A      m, 39

2473        A       A      m, 38

2475        A       A      m, 31

 480        A       A      m, 39

          Clinical
Isolate   diagnosis/        Other [ILLEGIBLE TEXT]
no.       source            isolated

2012      Endocarditis/     --
          blood culture

2410      Ulcus/lower       Staphylococc
          leg               aureus,
                            Streptococcus
                            pyogenes

2689      Puncture/hip      S. epidermidis
          joint             Corynebateri
                            pseudodiphth
                            icum

1682      Wound swab        S. aureus,
                            S. pyogenes

1935      Wound swab        S. aureus,
                            S. pyogenes

1836      Abscess/lower     S. aureus,
          leg               S. pyogenes

2661      Arthroscopy       Peptostreptoc
          aspirate/knee     prevotii, P. ma
          joint

2658      Fistula/hip       S. aureus
          joint

2674      Aspirate/hip      Streptococcus
          joint             group C

2670      Aspirate/knee     --
          joint

2667      Swab/hip joint    Coagulase-ne
                            staphylococci

2413      Ulcus/lower       S. aureus,
          leg               Streptococcus
                            group C/G

2446      Wound/upper       S. aureus, S.
          leg               pyogenes

2464      Ulcera/lower      Escherichia c
          arm               S. aureus, S.
                            pyogenes

2473      Ulcera/leg        E. coli,
                            Streptococcus
                            group C, G

2475      Ulcus/pretibial   Pseudomonas
                            aeruginosa,
                            Streptococcus
                            group C, G

  480     Ulcus/upper       S. aureus,
          leg               Clostridium
                            perfringens,
                            anaerobic flor

(a) Month/Year

(b) Paris, France; Hamburg, Germany; Zurich, Switzerland; Bern, Switzerland.

(c) DNA was isolated, digested with either EcoRI or PvuII, Electrophoresed, blotted, and probed fo [ILLEGIBLE TEXT] ribopatterns emerged, each with eight bands: patterns A, B, and C.

Many (10 [59%] of 17) C. diphtheriae isolates were from skin or subcutaneous [ILLEGIBLE TEXT] ulcers) in Swiss patients and were found with Staphylococcus aureus, [ILLEGIBLE TEXT] C/G streptococci, or (sometimes) with gram-negative rods. Most patients in this injecting drag users, homeless persons, prisoners, and (with one exception) men (30 to 58) years. Another subgroup (from Germany) consisted of six patients [ILLEGIBLE TEXT] infections (mentioned briefly in an earlier publication on coryneform bacteria [ILLEGIBLE TEXT] seventh patient had C diphtheriae biotype gravis [7]). The six had been [ILLEGIBLE TEXT] in Hamburg, which specializes in orthopedic surgery. Four had had implantation and had been admitted for prosthetic infections (with coagulase-negative [ILLEGIBLE TEXT] one pure culture of C. diphtheriae biotype mitis was obtained from a patient with one mixed culture with Peptostreptococcus magnus and Peptostreptococcuss [ILLEGIBLE TEXT] a patient who had a purulent knee infection after a fracture. Their average age [ILLEGIBLE TEXT] and, to our knowledge, none was a drag user. While the Hamburg isolates were [ILLEGIBLE TEXT] from every patient, their isolation dates stretched from March 1, 1995, until July isolated on the day of admission, and all of them were isolated by different [ILLEGIBLE TEXT] child (from France) had endocarditis with C. diphtheriae biotype mitis.

All isolates had identical antimicrobial susceptibility patterns. They were suscept (MIC, 0.25 [micro]/ml), amoxicillin-clavulanic acid (0.06 [micro]/ml), chloramphenicol (2 (0.25 [micro]/ml), clarithromycin (0.03 [micro]/ml), clindamycin (0.25 [micro]/ml), imipenem penicillin (0.25 [micro]/ml), and vancomycin (1 [micro]/ml). In contrast, all strains were [ILLEGIBLE TEXT] in the disk diffusion test (inhibition zone diameter 10 mm to 11 mm); their MICs doxycycline, and minocycline were 64 [micro]/ml, 16 [micro]/ml, and 16 [micro]/ml, [ILLEGIBLE TEXT] isolated tetracycline resistance was typical for the isolates from Swiss injecting [ILLEGIBLE TEXT] tetracycline resistance in nontoxigenic C. diphtheriae has otherwise very rarely [ILLEGIBLE TEXT] Europe (8). It has, however, been reported in toxigenic C. diphtheriae isolates [ILLEGIBLE TEXT] rarely, from Canada (9). The mechanism conferring this resistance in our strains investigated.

On analysis with restriction enzyme Pvu II, three different ribopatterns with [ILLEGIBLE TEXT] found among the strains (Figure). These patterns, though distinct, shared five (A (B vs. A) bands, respectively, and, therefore, may be considered related, as they pulsed-field gel electrophoresis were applied (10). This view is supported by the identical EcoRI patterns (not shown). All Swiss isolates were identical with the [ILLEGIBLE TEXT] whereas both the Swiss pattern and a second pattern were found among the Ham pattern was found exclusively in the single French strain. The staphylococcal and were not typed; they are no longer available to us.

[Figure ILLUSTRATION OMITTED]

Our isolates thus resemble those reported from Switzerland between 1990 and early 1996 (2,3), which were also often accompanied by S. aureus or betahemolytic streptococci. Such nontoxigenic C diphtheriae mitis may cause endocarditis, arthritis, and osteomyelitis (11,12). Most of the 52 isolates from France (11), examined with restriction enzymes different from ours and not available to us, also belonged to one ribotype. Their relatedness to our strains is unknown; however, they were largely tetracycline-susceptible. The two throat isolates from St. Petersburg, Russia, associated with a fatal diphtherialike disease (12) were not typed or tested for antibiotic susceptibility.

The origin of the isolates we describe is unknown. They may have been present ([ILLEGIBLE TEXT] the population for a long time. The mode of transmission is most likely common paraphernalia in the injecting drug use cases and in the endocarditis case; [ILLEGIBLE TEXT] to explain in the orthopedic infection subgroup. Although the Swiss and the Dani close, migration and contacts are not uncommon among injecting drug users. [ILLEGIBLE TEXT] have been distributed through the drugs themselves, as recently reported for S. [ILLEGIBLE TEXT] (13).

Our study may be representative for Switzerland and Germany, but since [ILLEGIBLE TEXT] strains to a central reference laboratory in these countries is not mandatory, we [ILLEGIBLE TEXT] frequency of these strains. These strains may also have spread to other European hypothesis can only be tested in a large multicenter study of European diphtheria

Acknowledgments

We thank J. Luthy-Hottenstein and V. Punter-Streit for excellent technical assistance and P. the strains from Hamburg. G. Funke is recipient of a European Society for Clinical Microbiology research fellowship.

References

(1.) Sloss JM, Hunjan RS. Incidence of non-toxigenic corynebacteria [ILLEGIBLE TEXT] personnel in Germany. J Infect 1996;33:139.

(2.) Gubler J, Huber-Schneider C, Gruner E, Altwegg M. An outbreak of non-[ILLEGIBLE TEXT] Corynebacterium diphtheriae infection: single bacterial clone causing [ILLEGIBLE TEXT] Swiss drug users. Clin Infect Dis 1998;27:1295-8.

(3.) Gruner E, Zuber PLF, Martinetti-Lucchini G, von Graevenitz A, Altwegg toxigenic Corynebacterium diphtheriae infections among Swiss intraveno[ILLEGIBLE TEXT] Medical Microbiology Letters 1992; 1:160-7.

(4.) Von Graevenitz A, Funke G. An identification scheme for rapidly and aero Gram-positive rods. Zentralbl Bakteriol 1996;284:246-54.

(5.) Martinetti-Lucchini G, Gruner E, Altwegg M. Rapid detection of [ILLEGIBLE TEXT] polymerase chain reaction. Medical Microbiology Letters 1992; 1:276-83.

(6.) National Committee for Clinical Laboratory Standards. Performance stand susceptibility testing; eighth informational supplement [NCCLS document (PA): The Committee; 1998.

(7.) von Graevenitz A, Frommelt L, Punter-Streit V, Funke G. Diversity of [ILLEGIBLE TEXT] infections following prosthetic joint insertion and open fractures. Infection

(8.) Patey O, Bimet F, Emond JP, Estrangin E, Riegel P, Halioua B, et al. Anti[ILLEGIBLE TEXT] of 38 non-toxigenic strains of Corynebacterium diphtheriae. J Antimicrob 1995;36:1108-10.

(9.) Rockhill RC, Sumarmo, Hadiputranto H, Siregar SP, Muslihun B. Tetracy Corynebacterium diphtheriae isolated from diphtheria patients in Jakarta, I Agents Chemother 1982;21:842-3.

(10.) Tenover FC, Arbeit RD, Goering RV, Mickelsen PA, Murray BE, Persing chromosomal DNA restriction patterns produced by pulsed-field gel [ILLEGIBLE TEXT] bacterial strain typing. J Clin Microbiol 1995;33:2233-9.

(11.) Patey O, Bimet F, Riegel P, Halioua B, Emond JP, Estrangin E, et al. [ILLEGIBLE TEXT] study of Corynebacterium diphtheriae systemic infections in France. J [ILLEGIBLE TEXT] 1997;35:441-5.

(12.) Rakhmanova AG, Lumio J, Groundstroem KWE, Taits BM, Zinserling V Fatal respiratory tract diphtheria apparently caused by nontoxigenic strains diphtheriae. Eur J Clin Microbiol Infect Dis 1997;16:816-20.

(13.) Streptokokken-Infektionen bei Drogensuchtigen in der Region Bern. Bulle fur Gesundheit 1997;44:3.

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Emerging Infectious Diseases Journal National Center for Infectious Diseases Centers for Disease Control and Prevention

URL: http://www.cdc.gov/ncidod/eid/vol5no3/funke.htm

Guido Funke,(*) Martin Altwegg,(*) Lars Frommelt,([dagger]) and Alexander von Grae

(*) University of Zurich, Zurich, Switzerland; and ([dagger])Endo-Klinik, Hamburg, Germa

Dr. Funke is director of clinical microbiology at Gartner & Colleagues Laboratories in Wein of expertise are clinical and systematic bacteriology. Research interests include taxonomy and [ILLEGIBLE TEXT] coryneform bacteria and actinomycetes, as well as rapid methods for identification and [ILLEGIBLE TEXT] relevant bacteria.

Address for correspondence: Alexander von Graevenitz, Department of Medical Microbiolog Gloriastra sse 32, CH-8028 Zurich, Switzerland; fax: [phone]; e-mail: [e-mail].c