Surface spreading motility shown by a group of phylogenetically related, rapidly growing pigmented mycobacteria suggests that motility is a common property of mycobacterial species but is restricted to smooth colonies

Abstract

Motility in mycobacteria was described for the first time in 1999. It was reported that Mycobacterium smegmatis and Mycobacterium avium could spread on the surface of solid growth medium by a sliding mechanism and that the presence of cell wall glycopeptidolipids was essential for motility. We recently reported that Mycobacterium vaccae can also spread on growth medium surfaces; however, only smooth colonies presented this property. Smooth colonies of M. vaccae do not produce glycopeptidolipids but contain a saturated polyester that is absent in rough colonies. Here, we demonstrate that Mycobacterium chubuense, Mycobacterium gilvum, Mycobacterium obuense, and Mycobacterium parafortuitum, which are phylogenetically related to M. vaccae, are also motile. Such motility is restricted to smooth colonies, since natural rough mutants are nonmotile. Thin-layer chromatography analysis of the content of cell wall lipids confirmed the absence of glycopeptidolipids. However, compounds like the above-mentioned M. vaccae polyester were detected in all the strains but only in smooth colonies. Scanning electron microscopy showed great differences in the arrangement of the cells between smooth and rough colonies. The data obtained suggest that motility is a common property of environmental mycobacteria, and this capacity correlates with the smooth colonial morphotype. The species studied in this work do not contain glycopeptidolipids, so cell wall compounds or extracellular materials other than glycopeptidolipids are implicated in mycobacterial motility. Furthermore, both smooth motile and rough nonmotile variants formed biofilms on glass and polystyrene surfaces.

FIG. 1.

Smooth and rough colonies of M. chubuense (A), M. gilvum (B), M. obuense (C), M. parafortuitum (D), and M. vaccae (E). The images are representative of hundreds of colonies obtained through the study.

FIG. 2.

SEM images of smooth colonies of M. chubuense (A), M. gilvum (B), M. obuense (C), M. parafortuitum (D), and M. vaccae (E). The right column shows the same sample as the left column but at greater magnification. These images are representative of the studies performed with six colonies of each species.

FIG. 3.

SEM images of rough colonies of M. chubuense (A), M. gilvum (B), M. obuense (C), M. parafortuitum (D), and M. vaccae (E). The right column shows the same sample as the left column but at greater magnification. These images are representative of the studies performed with six colonies of each species.

FIG. 4.

Spreading of smooth colonies on motility medium after 6 days of growth. (A) M. chubuense; (B) M. gilvum; (C) M. obuense; (D) M. parafortuitum; (E) M. vaccae. Arrows indicate the external margins of the colonies. The right column shows pictures obtained with a binocular stereomicroscopy. These images are representative of the studies performed with 10 colonies of each species.

FIG. 5.

Spreading of rough colonies on motility medium after 6 days of growth. (A) M. chubuense; (B) M. gilvum; (C) M. obuense; (D) M. parafortuitum; (E) M. vaccae. Arrows indicate the external margins of the colonies. In the right column are pictures obtained with a binocular stereomicroscopy. These images are representative of the studies performed with 10 colonies of each species.

FIG. 6.

TLC images of chloroform-methanol extracts from M. vaccae (lanes 1 and 2), M. gilvum (lanes 3 and 4), M. obuense (lanes 5 and 6), M. chubuense (lanes 7 and 8), and M. parafortuitum (lanes 9 and 10). In the odd-numbered lanes, extracts are from smooth colonies, and in the even-numbered lanes, extracts are from rough colonies. TLC was developed with methanol-chloroform (90:10, vol/vol) and revealed with anthrone. Red spots indicate SP and SP-L.

FIG. 7.

Cellular adhesion of smooth and rough variants of M. chubuense (A), M. gilvum (B), M. obuense (C), M. parafortuitum (D), and M. vaccae (E) strains to glass tubes at 1, 2, 3, 4, and 7 days of incubation. The cellular adhesion level is expressed as the number of CFU/cm². The results are expressed as means ± standard deviations obtained from triplicates. The results are representative of one out of three independent experiments. Cellular adhesion levels of smooth variants were significantly higher than those of rough variants (*, P < 0.05; **, P < 0.01). Cellular adhesion levels of rough variants were significantly higher than those of smooth variants (&, P < 0.05).

FIG. 8.

Cellular adhesion of smooth and rough variants of M. chubuense (A), M. gilvum (B), M. obuense (C), M. parafortuitum (D), and M. vaccae (E) strains to polystyrene tubes at 1, 2, 3, 4, and 7 days of incubation. The cellular adhesion level is expressed as the number of CFU/cm². The results are expressed as means ± standard deviations obtained from triplicates. The results are representative of one out of three independent experiments. Cellular adhesion levels of smooth variants were significantly higher than those of rough variants (**, P < 0.01). Cellular adhesion levels of rough variants were significantly higher than those of smooth variants (&&, P < 0.05).