First Case of Mycobacterium longobardum Infection
A 71-yr-old man presented with symptoms of pain, redness, and swelling in the left elbow on May 7, 2012. The patient had been previously diagnosed with left elbow bursitis and had undergone surgery to resect the infected tissue at a university hospital in January 2011. At that time, a pathology review of the excised tissue confirmed the diagnosis of tuberculous bursitis with chronic granulomatous inflammation, without isolation of Mycobacterium tuberculosis (MTB). The patient received the antituberculosis drugs of isoniazid, rifampicin, ethambutol, and pyrazinamide for seven months.\nThe patient had a history of being treated for hypertension and diabetes. Physical examination at presentation revealed surgical scarring and tenderness around the left elbow. The patient's electrolytes, liver function, and complete blood counts were normal. His blood pressure was 107/76 mmHg and his two-hour postprandial glucose level was 106 mg/dL. Magnetic resonance imaging (MRI) of the left elbow revealed localized bone marrow signal alterations at the left ulnar olecranon and substantial effusion with marked synovial hypertrophy in the radiocapitellar joint.\nThe patient was assessed as having tuberculous osteomyelitis according to the history and MRI results. The left elbow was incised and drained using sterilized devices on May 8, 2012. Histological examination of the resected tissue showed necrotizing granulomatous inflammation (). No organisms were detected on Gram or Ziehl-Neelsen staining, and the direct culture of the tissue sample for bacteria was sterile. In addition, there was no evidence of malignancy. The biopsied tissue was cultured for mycobacteria, and mycobacterial growth was observed on Löwenstein-Jensen medium 26 days later. The isolate was acid-fast bacillus and colonies grew rough and unpigmented both in the dark and after light exposure. The mycobacterium was identified as NTM using the Seeplex MTB/NTM ACE Detection Kit (Seegene Inc., Seoul, Korea).\nThe 16S rRNA gene was sequenced for the NTM identification. The primers and PCR setting used to amplify the target region, the first 500 bp of the 16S rRNA gene, were as previously described [, ]. Sequences were analyzed using an ABI PRISM 3730 series DNA Analyzer (Applied Biosystems, Foster City, CA, USA). For the first 500 bp of the 16S rRNA gene sequence, the isolate showed a 98.6% identity match with GenBank sequence JN571166.1 (M. longobardum), a 97.7% identity match with GenBank sequence FJ268583.1 (M. senuense), and a 97.5% identity match with GenBank sequences NR_043905.1 (M. senuense) and GQ184162.1 (M. terrae). However, the Clinical and Laboratory Standards Institute guidelines indicate that a Mycobacterium sp. that has a 95-99% nucleotide identity with 16S rRNA gene sequences of other species cannot be definitively identified by 16S rRNA gene sequencing [].\nTortoli et al. [] reported that 65-kDa heat shock protein (hsp65) and full 16S rRNA gene sequencing are useful for the identification of M. longobardum. For accurate identification, the hsp65 and additional 16S rRNA genes of the isolate were analyzed. Using primers amplifying 468-1,053 bp (F1 5'-TGC CAG CAG CCG CGG TAA-3', R1 5'-CGG GAC TTA ACC CAA CAT CT-3') and 1,010-1,500 bp (F2 5'-TGG CTG TCG TCA GCT CGT-3', R2 5'-AAG GAG GTG ATC CAA CCG CA-3') of the 16S rRNA gene, PCR was performed as described for sequencing of the first 500 bp of the 16S rRNA gene sequence. For the full 16S rRNA gene sequence, the isolate showed a 99.2% identity match with GenBank sequence JN571166.1 (M. longobardum), a 98.6% identity match with GenBank sequence HM770865.1 (M. terrae), and a 98.4% identity match with GenBank sequence GU084182.2 (M. heraklionense). The 16S rRNA gene phylogenetic tree of the mycobacterium isolated in this case was most closely related to JN571166.1 (M. longobardum) and was classified as M. longobardum (). The primers and PCR settings used to amplify the hsp65 gene were as described previously []. Moreover, hsp65 sequencing of the isolate showed a 99.4% match with sequences JN571166.1 (M. longobardum), EF601223.1 (M. terrae), and JX154097.1 (M. terrae). We concluded that the isolate was M. longobardum.\nAfter identifying M. longobardum, the patient was prescribed clarithromycin and ethambutol empirically. The antimicrobial susceptibility test (AST) using the broth microdilution method revealed that the isolate was susceptible to clarithromycin, ethambutol, moxifloxacin, and sulfamethoxazole, but resistant to amikacin, ciprofloxacin, and rifampicin. The therapy was maintained, and the patient was cured without further complications.