The Mycococcus Form of Mycobacteria

Abstract

SUMMARYCultures of form 2 mycobacteria, which had been recently isolated from form 1 of strains of Mycobacterium tuberculosis and atypical mycobacteria, were cultivated on nutrient agar in screw-capped bottles and kept unopened for 6 weeks at room temperature. Gram-positive granules developed in organisms which had penetrated into the medium. When these organisms were incubated in digest broth at 37' with intermittent aeration, the granules were liberated into the medium and gradually increased in size to that of a small coccus. After a resting period of about 2 weeks, the ' cocci ' started to multiply independently, forming diplococci and tetrads.At this stage the cocci yielded colonies on subculture on nutrient agar. The cocci were identified as members of the genus Mycococcus, family Mycobacteriaceae, order Mycobacteriales (Krassilnikov, 1959). Mycococci were not obtained in broth cultures which were either not aerated or were aerated frequently or continuously. Prolonged subcultivation of the parent form 2 strains on nutrient agar prevented the development of mycococci in broth. Evidence, such as the failure to isolate mycococci from uninoculated but aerated media, is submitted that the mycococci were not contaminants. Previous work on isolation of similar cocci from different members of the order Actinomycetales, including human and bovine strains of M . tuberculosis is reviewed.

Cell Wall Deficiency in Mycobacteria: Latency and Persistence

“…Sometimes, persistent M. tuberculosis bacteria bearing cell wall alterations may remain undetected by the classic Ziehl-Neelsen staining (Seiler et al, 2003). Appearance of polymorphic non-acid fast forms and coccoids in cultures of mycobacteria has been observed by other authors (Chandrasekhar & Ratnam , 1992;Csillag, 1964;Juhasz, 1962;Miller, 1932;Xalabarder, 1958). Morphological forms of different sizes and shapes (short coccobacilli and long rods, oval or round coccoid cells, large spherical bodies and giant filaments) in mycobacterial L-form cultures obtained after starvation stress, were observed by us with scanning electron microscopy ( Fig.4, n. d.).…”

“…Light and electron microscopy also provided interesting results about the appearance of non-acid fast coccoid cell morphology of stressed M. tuberculosis, that support observations of other authors. The appearance of non-acid fast coccoids in cultures of mycobacteria has been reported by others in the beginning of the last century but the phenomenon was not clearly explained and proven at that time (Csillag, 1964;Juhasz, 1962;Xalabander, 1958;). More surprising was the fact that mycobacterial coccoid L-forms not only mimicked the morphology of staphylococci or other coccus-shaped bacteria, but also exhibited extremely rapid growth and colonial development in contrast to classical TB bacilli (n. d. Standard plating techniques are often inadequate for accurate enumeration of microbial dormant forms, because some of them may be in a "nonculturable" state (Shleeva et al, 2010).…”

“…Coccoid forms of mycobacteria, called "mycococcus", were obtained in vitro by Csillag in 1964. Mycococcii were grown from M. tuberculosis and were similar to the morphology of staphylococci (Csillag, 1964). Genetic analysis of mycobacterial coccoids however, performed by us through amplification of 16SrRNA gene fragment, 16S-23S rRN gene Internal Transcribed Spacer sequences and IS 6110 PCR, verified them as M. tuberculosis (n. d.).…”

“…tuberculosis is known to exhibit extreme pleomorphism in certain circumstances. Various morphological forms of mycobacteria were observed by many authors and were described as "mycococcus form" (Csillag, 1964), large "amoeba-like cells" (Imaeda, 1975), giant noncellular structures or so called "budding yeast-like structures" (Koch, 2003), "elementary bodies and filament structures" (Merkal et al, 1973) "endospores" (Ghosh et al,2009;Traag et al, 2010) and "ovoid cells" ( Shleeva et al, 2011). Mycobacteria are unique among procaryotes with their cell wall structure, containing tightly packed mycolic acids that provide TB bacilli with efficient protection and remarkable capacity to resist to various exogenous stress conditions.…”

Morphological Characterization of Mycobacterium tuberculosis

“…But, 12 weeks after INH and PZA treatment was withdrawn, one-third of the mice developed full-blown active TB, with nearly two-thirds displaying disease after 24 weeks. Csillag (1962Csillag ( , 1963Csillag ( , and 1964 considered Mycobacteria as dimorphic organisms in the same sense as are some pathogenic fungi, for instance, Histoplasma capsulatum. The usual acid fast form of the mycobacteria was termed 'form I' and the form which was not acid fast was termed 'form 2'.…”