Spheroplastic phase of mycobacteria isolated from patients with Crohn's disease

Abstract

Two strains of an unclassified Mycobacterium species were isolated after 18 and 30 months of incubation of media inoculated with resected intestinal tissues from patients with Crohn's disease. These strains represented the third and fourth isolates of this organism from Crohn's disease patients. Ultrastructural examination of this strain and two previously isolated strains revealed the presence of spheroplasts which eventually transformed into the bacillary form of a previously unrecognized Mycobacterium species. These cell wall-deficient forms did not stain with conventional dyes and failed to grow on hypertonic media. Restriction polymorphism of the ribosomal DNA genes was used to determine the relationship between the cell wall-deficient and baciliary forms. Identical restriction patterns of the ribosomal DNA genes were found between the spheroplasts and Mycobacterium sp. isolates with EcoRI, BamHI, and XhoI restriction endonucleases, thus providing definitive evidence of their origin. Unidentified spheroplasts were isolated from an additional 12 patients with Crohn's disease, of which 7 of 10 seroagglutinated with antiserum prepared against the Mycobacterium sp. Spheroplasts were isolated from 16 of 26 (61%) patients with Crohn's disease but not from tissues of 13 patients with ulcerative colitis or 13 patients with other diseases of the bowel. These findings support the role of mycobacteria as etiologic agents in some cases of Crohn's disease.

Lack of support for a common etiology in Johne's disease of animals and Crohn's disease in humans

“…Those who support the hypothesis that M. paratuberculosis is responsible for CD reconcile their failures at classic recovery by culture by reporting that they often find small translucent colonies of 0.5 mm size, on the Herrold's egg yolk media, after many months of incubation, which when stained by an acid-fast method contain what are described as coccoid bodies and other amorphous material, pleomorphic forms and rods, or spheroplast-like agents (36)(37)(38)(39)(40). Some of these, one-third it is claimed, when tested by PCR for IS 900 were identified as M. paratuberculosis (41,42).…”

Zoonotic potential ofMycobacterium aviumssp.paratuberculosis: the current position

“…Consequently, the primary source of infection with Map is an infected animal. There is evidence that Map can exist in vegetative, cell wall deficient (Chiodini et al. 1986), and dormant (Whittington et al.…”

“…The three strains, designated Linda, Ben and Dominic, had taken up to 18 months for primary isolation. It was later reported that all three strains had originally been isolated as nonacid‐fast coccobacillary forms that had the ultrastructural appearance of sphaeroplasts (cell wall deficient forms) which, after several months incubation, transformed into characteristic Map ‐like organisms (Chiodini et al. 1986).…”

Evaluation of Surgical Tissue From Patients with Crohnʼs Disease for the Presence of Mycobacterium avium Subspecies paratuberculosis DNA by In Situ Hybridization and Nested Polymerase Chain Reaction

“…The spheroplast cultures then reverted to acid-fast Mycobacterium-like organism after a single in vitro passage. 6,7 They were confirmed as MAP culture using IS900 probe. 8 This was followed by several studies, which resulted in mixed results in which MAP or MAP DNA presence ranged from 0 to 100%.…”