Characterization of rough and smooth morphotypes of Mycobacterium abscessus isolates from clinical specimens

Abstract

Mycobacterium abscessus, which consists of the two subspecies M. abscessus subspecies abscessus and M. abscessus subspecies bolletii, can produce rough or smooth colony morphologies. Here we analyzed 50 M. abscessus isolates cultured from the respiratory specimens of 34 patients, 28 (82%) of whom had cystic fibrosis (CF), with respect to their colony morphologies and antibiotic susceptibilities. The overall proportions of occurrences of the two morphotypes were similar, with specimens from 50% of the patients showing a rough and 38% showing a smooth morphotype. A total of 12% of the specimens from the patients showed both morphotypes simultaneously. At the subspecies level, the proportions of rough and smooth morphotypes differed substantially; 88% of rough morphotypes belonged to M. abscessus subspecies abscessus, and 85% of smooth morphotypes belonged M. abscessus subspecies bolletii. Inducible clarithromycin resistance due to the Erm(41) methylase, as well as high-level resistance to clarithromycin due to mutations within the rrl gene, occurred independently of the morphotype. The MIC50s of amikacin and cefoxitin were identical for the two morphotypes, whereas the MIC50s of tigecycline were 0.25 μg/ml for the rough morphotype and 2.0 μg/ml for the smooth morphotype. Our results show that the smooth morphotype was more dominant in respiratory specimens from CF patients than previously thought. With respect to resistance, colony morphology did not affect the susceptibility of Mycobacterium abscessus to the first-line antibiotics clarithromycin, amikacin, and cefoxitin.

FIG 1

Growth characteristics of rough and smooth phenotypes on 7H11 agar cultured at 37°C: representative single rough (left) and smooth (right) colonies.

FIG 2

Colony morphology of the primary subculture and results of pulsed-field gel electrophoresis of rough and smooth strains from the four patients (P2, P17, P31, and P37) that produced a mixed phenotype. Subcultures from primary specimens that had been kept as frozen stocks were recultured on 7H11 agar at 37°C, and photos were taken. Two patients (P2 and P17) had predominantly rough morphotypes, and two patients (P31 and P37) had predominantly smooth morphotypes. From each patient, one smooth colony and one rough colony were subcultured until the morphotypes remained stable before genomic DNA was prepared, XbaI digested, and separated by pulsed-field gel electrophoresis. The gel photo shows a comparison of the smooth (s) and rough (r) morphotypes from each of the four patients (P2, P17, P31, and P37). M, molecular marker; ATCC, M. abscessus ATCC 19977 type strain.