Correspondence

Mycobacterium ulcerans and Osteomyelitis

N Engl J Med 1993; 329:582-583August 19, 1993

Article

To the Editor:

In their report in the April 8 issue, Hofer et al. describe disseminated osteomyelitis due to Mycobacterium ulcerans infection after a snakebite1. We do not believe that the authors have unambiguously established M. ulcerans as the infecting organism.

M. haemophilum has been reported to cause lymphadenitis and skin lesions in immunocompetent patients. In immunocompromised patients, disseminated infections involving the skin, subcutaneous tissues, bones, and joints have been described2-4.

Culture medium supplemented with iron is essential for growing M. haemophilum in vitro, and this requirement is characteristic of the species. Neither medium used by Hofer et al. (Lowenstein-Jensen or Middlebrook 13A) contains iron, and consequently neither would have supported the growth of M. haemophilum.

To substantiate the diagnosis of M. ulcerans, the authors determined a 15-bp nucleotide sequence within the 16S ribosomal DNA (rDNA) gene of isolates of M. marinum and M. ulcerans and compared these sequences with those of amplified DNA from the patient's infected tissues. The latter sequences matched those of M. ulcerans. The reference species differed only at nucleotide 1248, a finding that the authors state was in keeping with previous data indicating 99.9 percent homology in the 16S rDNA gene sequence of these species. However, neither of the two references cited5,6 mentions M. ulcerans, although both include data on M. marinum and other mycobacteria.

A search of the data bases of GenBank (release 76) and the European Molecular Biology Laboratory ([EMBL] version 34) reveals two entries for the 16S rDNA gene of M. ulcerans, both unpublished (Entrez: Sequences, National Library of Medicine, Bethesda, Md.). These data confirm the nucleotide sequences presented by Hofer et al., but there are also several other differences in sequence between the species, including two in regions adjacent to the sequences shown in the authors' Figure 2. The same data bases include no entry for M. haemophilum. In the absence of specific information about sequence, the relatedness of M. haemophilum to either M. ulcerans or M. marinum at the level of rDNA sequence remains speculative.

The clinical and microbiologic observations presented are compatible with M. haemophilum infection. The data on nucleotide sequence that are currently available do not rule out this possibility.

Mar Kristjansson, M.D.
Robert D. Arbeit, M.D.
Veterans Affairs Medical Center, Boston, MA 02130

6 References

1. 1

   Hofer M, Hirschel B, Kirschner P, et al. Disseminated osteomyelitis from Mycobacterium ulcerans after a snakebite. N Engl J Med 1993;328:1007-1009
   Full Text | Web of Science | Medline

2. 2

   McBride ME, Rudolph AH, Tschen JA, et al. Diagnostic and therapeutic considerations for cutaneous Mycobacterium haemophilum infections. Arch Dermatol 1991;127:276-277
   CrossRef | Web of Science | Medline

3. 3

   Kristjansson M, Bieluch VM, Byeff PD. Mycobacterium haemophilum infection in immunocompromised patients: case report and review of the literature. Rev Infect Dis 1991;13:906-910
   CrossRef | Medline

4. 4

   Yarrish RL, Shay W, LaBombardi VJ, Meyerson M, Miller DK, Larone D. Osteomyelitis caused by Mycobacterium haemophilum: successful therapy in two patients with AIDS. AIDS 1992;6:557-561
   CrossRef | Web of Science | Medline

5. 5

   Rogall T, Wolters J, Flohr T, Bottger EC. Towards a phylogeny and definition of species at the molecular level within the genus Mycobacterium. Int J Syst Bacteriol 1990;40:323-330
   CrossRef | Medline

6. 6

   Rogall T, Flohr T, Bottger EC. Differentiation of Mycobacterium species by direct sequencing of amplified DNA. J Gen Microbiol 1990;136:1915-1920
   Medline

To the Editor:

Hofer et al. suspected that the M. ulcerans infection in their patient was caused by a snakebite, but they did not provide evidence to support their suspicion. First, what was the evidence that a snakebite occurred? Was the bite witnessed by someone other than the three-year-old patient? Was a snake captured or identified? An unverified report of a snakebite is not sufficient evidence. Almost 10 percent of reported snakebites turn out not to be snakebites on further investigation1. Second, if a bite occurred, what was the evidence that the M. ulcerans infection started with the bite? Did clinical characteristics typical of an M. ulcerans infection develop in the bite wound? Third, if a bite occurred, and if the wound was infected with M. ulcerans, what was the evidence that the wound was not inoculated with bacteria after the bite? Plants are sometimes applied to snakebite wounds as a nostrum against envenomation, and M. ulcerans infections have been associated with certain types of plants2,3. The lesions associated with the disseminated M. ulcerans infection apparently did not appear until several months after the bite. Therefore, the infection might not have occurred until several months after the bite. M. ulcerans might have contaminated one of the surgical wounds, or it might have disseminated from the lungs after inhalation4. From the evidence presented, the connection between the alleged snakebite and the M. ulcerans infection is quite tenuous.

Harrison G. Weed, M.D.
Ohio State University College of Medicine, Columbus, OH 43210

4 References

1. 1

   Weed HG. Nonvenomous snakebite in Massachusetts: prophylactic antibiotics are unnecessary. Ann Emerg Med 1993;22:220-224
   CrossRef | Web of Science | Medline

2. 2

   Looareesuwan S, Viravan C, Warrell DA. Factors contributing to fatal snake bite in the rural tropics: analysis of 46 cases in Thailand. Trans R Soc Trop Med Hyg 1988;82:930-934
   CrossRef | Web of Science | Medline

3. 3

   Barker DJP. Epidemiology of Mycobacterium ulcerans infection. Trans R Soc Trop Med Hyg 1973;67:43-50
   CrossRef | Web of Science | Medline

4. 4

   Hayman J. Postulated epidemiology of Mycobacterium ulcerans infection. Int J Epidemiol 1991;20:1093-1098
   CrossRef | Web of Science | Medline

Author/Editor Response

The authors reply:

To the Editor: We attempted to culture the microorganism on iron-supplemented medium, without success. Because there was no growth on any medium we tried, we extracted DNA from biopsy specimens containing acid-fast bacilli. We identified the probable cause of the disseminated osteomyelitis as M. ulcerans on the basis of the sequence of 16S rRNA. Species of mycobacteria usually differ in the hypervariable regions of the 16S rRNA gene. This gene contains specific signature sequences that clearly distinguish M. ulcerans or M. marinum from M. haemophilum1. In addition to the stretch of 15 nucleotides that we reported, where a single base change differentiates M. marinum (EMBL accession number X 52920) from M. ulcerans (EMBL accession number X 58954), we have sequenced several hundred other nucleotides, including the hypervariable regions. At each relevant location, the sequence of the DNA extracted from specimens from our patient was characteristic of that of M. ulcerans and differed from that of M. haemophilum1.

Weed raises questions about the history of snakebite. Because the child was transferred alone from Africa to our hospital, it was not possible to ascertain the history, as we would have liked, and as Weed suggests. Therefore, the link between the multifocal osteomyelitis and the snakebite remains somewhat tenuous. It is, however, intriguing. The natural reservoir of M. ulcerans is unknown, despite attempts to find the pathogen in plants and domestic animals in Africa2. Snakes' mouths have not been checked.

B. Hirschel, M.D.
Hopital Cantonal Universitaire, CH-1211 Geneva 14, Switzerland

E. Bottger, M.D.
Medizinische Hochschule Hannover, D-3000 Hannover, Germany

2 References

1. 1

   Kirschner P, Meyer A, Bottger EC. Genotypic identification and detection of mycobacteria: facing novel and uncultured pathogens. In: Persing DT, White TD, Smith TF, eds. Diagnostic molecular microbiology. Washington, D.C.: American Society for Microbiology, 1993:173-90.
   

2. 2

   Portaels F. Mycobacterioses. In: Janssens PG, Kivits M, Vuyelsteke J, eds. Medecine et hygiene en Afrique Centrale de 1885 a nos jours. Vol. 2. Brussels, Belgium: Fondation du Roi Baudoin, 1992.