Crohn's disease is a chronic granulomatous ileocolitis, of unknown etiology, which generally affects the patient during the prime of life. Medical treatment is supportive at best and patients afflicted with this disorder generally live with chronic pain, in and out of hospitals, throughout their lives. The disease bears the name of the investigator that convincingly distinguished this disease from intestinal tuberculosis in 1932. This distinction was not universally accepted and the notion of a mycobacterial etiology has never been fully dismissed. Nevertheless, it was 46 years after the distinction of Crohn's disease and intestinal tuberculosis before research attempting to reassociate mycobacteria and Crohn's disease was published. Recently, there has been a surge of interest in the possible association of mycobacteria and Crohn's disease due largely to the isolation of genetically identical pathogenic M. paratuberculosis from several patients with Crohn's disease in the United States, the Netherlands, Australia, and France. These pathogenic organisms have been isolated from only a few patients and direct evidence for their involvement in the disease process is not clear; however, M. paratuberculosis is an obligate intracellular organism and strict pathogen which strongly suggests some etiologic role. Immunologic evidence of a mycobacterial etiology, as assessed by humoral immune determinations, have been conflicting, but evaluation of the more relevant cellular immunity have not been performed. Data from histochemical searches for mycobacteria in Crohn's disease tissues have been equally conflicting with acid-fast bacilli detected in 0 to 35% of patients. Animal model studies have demonstrated the pathogenic potential of isolates as well as elucidating the complexity of mycobacteria-intestinal interactions. Treatment of Crohn's disease patients with anti-mycobacterial agent has not been fully assessed, although case reports suggest efficacy. The similarities in the pathology, epidemiology, and chemotherapy of Crohn's disease and the mycobacterioses are discussed. The issue is froth with controversy and the data generated on the association of mycobacteria and Crohn's disease are in their infantile stages such that a general conclusion of the legitimacy of this association cannot be made. While no firm evidence clearly implicates mycobacteria as an etiologic agent of Crohn's disease, the notion is supported by suggestive and circumstantial evidence, and a remarkable similarities to other known mycobacterial diseases.

The disease that Crohn, Ginzberg, and Oppenheimer described in 1932 was a chronic low-grade inflammation of the terminal ileum (59). Earlier cases may have been documented (196, 298), but the authors failed to receive recognition for describing a new disorder. These earlier cases were called nonspecific granulomata and were sorted from those that had previously been termed hyperplastic tuberculosis of the intestine. Although Crohn's disease was first described as a segmental disease of the small intestine, in 1960, it was recognized that the same disorder affected the colon and had been confused with ulcerative colitis (161). In recent years the lesions of Crohn's disease have been recognized in the mouth, larynx, esophagus, stomach, skin, muscle, synovial tissue, and bone (17, 141, 142, 152, 171, 182, 206, 297). Thus, Crohn's disease may be considered a newly recognized disease, with a defined clinical and pathologic description dating back only to the 1960's. Although the terms Crohn's disease, Crohn's colitis, Crohn's ileitis, and regional ileitis have been with us longer, there is uncertainty as to the accuracy of these diagnoses prior to 1960. To this date, Crohn's disease and ulcerative colitis continue to be confused clinically and the term inflammatory bowel disease (IBD) was developed to comprise both diseases.

Patients afflicted with this disorder generally suffer with chronic weight loss, abdominal pain, diarrhea or constipation (obstruction), vomiting, and generalized malaise. Between 70 to 80% of Crohn's disease patients require surgical resection of the diseased intestine (83, 102, 120, 243). Difficulties usually are not ended by surgical intervention, and most patients will suffer recurrences and require further surgical procedures (71, 109, 123, 161, 181, 291). Generally, patients live with chronic pain, in and out of hospitals, throughout their lives. Mortality is approximately 6% (83, 239). Perhaps more important than mortality, is the quality of life of Crohn's disease patients. Less than 50% of patients consider their quality of life to be "good"; suboptimal psychosocial function is recorded in 30-54% of patients (83, 230, 291). These patients with inflammatory bowel disease (an estimated 2 million, 200,000 whom are children, in the United States alone), represent a very unhappy population with little prospect for relief. The etiology remains obscure and medical treatment is supportive at best (102, 238, 243). Twenty-five years after the original description, Crohn and Yarnis wrote (61): "From this small beginning we have witnessed the evolution of a Frankenstein monster that, if not threatening to life, frequently results in serious illness, often prolonged and debilitating."

........ To this date, the etiology of Crohn's disease, being infectious, transmissible, or not, has eluded the scientific community. Now, over 55 years since its distinction from tuberculosis, attention has been driven back to the beginning: "Is Crohn's disease a mycobacterial disease after all?" (97).

Medical historians suggest that Crohn's disease may first have been described as early as 1682-1771, or even earlier (143). Reports of diseases suggestive of Crohn's disease have appeared in 1806, 1813, 1828, 1875, 1907, 1908, 1909, and 1913 (143). Whether these cases actually were Crohn's disease, will remain unknown. Mycobacteria were not discovered until 1874 ....

...... The landmark article by Crohn, Ginzberg, and Oppenheimer (59) recognized regional ileitis as a separate and unique disease entity and displaced the long-held belief of a mycobacterial etiology. We now know that hypertrophic intestinal tuberculosis (3) and tuberculosis without caseation or demonstrable acid-fast bacilli (296) do exist, as well as a distinct disease known as Crohn's disease. Nevertheless, over the years, the notion recurs that Crohn's disease might in fact be mycobacterial in origin.

Attempts to isolate mycobacteria from Crohn's disease patients dates back before the time that this disease was recognized as a distinct entity.

Perhaps the first concerted effort to isolate mycobacteria from Crohn's disease patients was presented by Van Patter, W. (Ph.D. thesis, University of Minnesota, 1952). In these studies, Van Patter reported the results of 1,762 cultures from 43 patients with Crohn's disease. By employing 7 different types of media and incubation periods up to 15 months for some cultures, he isolated acid-fast organisms from 3 patients (7%) after 6, 7.5, and 8 months incubation. These organisms could not be subcultured and were never formally identified.

For the next 25 years, not a single report appeared on the attempt to isolate mycobacteria from Crohn's disease patients. Although undoubtedly attempts were made over the years, ....

.... In 1978, a revival of the notion that mycobacteria might be related to Crohn's disease occurred with the articles by Burnham et al (33, 34). These authors described the isolation of M. kansasii from the lymph node of a single patient with Crohn's disease and pleomorphic material, suggestive of cell wall deficient (CWD) organisms, from 22 of 27 Crohn's disease patients, 7 of 13 ulcerative colitis patients, and 1 of 11 controls. ....

.... The efforts of Stanford et al were reviewed and up dated at a recent symposium (266). Since 1974, these investigators have examined over 200 surgical specimens and have isolated pleomorphic, variable acid-fast, organisms from 42 of 76 (55%) Crohn's disease patients, 17 of 27 (52%) ulcerative colitis patients, and 3 of 41 (7%) controls. The organisms remain unidentified although efforts are in progress to classify them more precisely. ....

..... In 1984 there was again a surge of activity on the role of mycobacteria and Crohn's disease, and yet a different Mycobacterium species. From this period on, there have been more reports on mycobacteria and Crohn's disease than in the last 50 years, ....

.... The authors concluded that their isolates were strains of M. paratuberculosis or a biovariant of that species and suggested that this organism plays an etiologic role in at least some cases of Crohn's disease. An editorial that accompanied some of these papers (97) suggested that these studies "provide the most intriguing evidence yet generated regarding a possible cause of this important illness" and that "scientists have come closer than ever to fulfilling Koch's postulates and developing a test system for Crohn's disease". ....

..... Some of these CWD forms required up to 1-1/2 years incubation for primary emergence of colonies. ....

.... Based on available information about mycobacterial spheroplasts, which suggests that only bacillary forms are pathogenic, they postulated that a very slow rate of reversion with subsequent local hypersensitivity-type immunologic responses could account for the chronicity of Crohn's disease. ....

.... In their article, Yoshimura et al (307) presented additional data supporting the identification of the Crohn's disease-isolated mycobacteria previously reported (48) as M. paratuberculosis.

The culture results of Graham and co-workers (106) provide some useful and important information and illustrate the ubiquitousness of some Mycobacterium spp. ....

.... An interesting feature of the culture results of Graham et al (106) is the specimen type from which mycobacteria were isolated. Except for a single strain of M. fortuitum complex, mycobacteria could not be isolated from resected tissues; but 35 strains of mycobacteria (predominantly MAI and M. fortuitum) were isolated from biopsy specimens of aphthous ulcers. These ulcers provide a suitable micro-environment for the propagation of such environmental organisms. On the other hand, Graham et al (106) isolated as yet unidentified spheroplasts primarily from resected tissues of Crohn's disease patients rather than aphthous ulcers. Retrospectively, these culture data appear to support, rather than refute, a CWD mycobacterial etiology. Thus, as in all other diseases, the area from which material is obtained for culture is of great importance, as are the techniques applied to tissue processing.

..... S. R. Pattyn, F. Portaels, and Y. Van Maercke presented their culture results at the meeting of the International Working Group on Mycobacterial Taxonomy (IWGMT) held in Bithoven, The Netherlands in September of 1987 .... These workers examined tissues from 32 patients with Crohn's disease and demonstrated acid fast bacilli in 11 (34%) by acid-fast staining. Cultivation attempts yielded 2 strains of M. chelonei which were said to be mycobactin-dependent; mycobacteria could not be isolated from the remaining 9 cases in which acid-fast bacilli were observed. The authors acknowledged that their processing technique, i.e., 0.15% benzalkonium chloride and 0.5% NaOH, may have been too deleterious for recovery of other acid-fast bacilli; M. paratuberculosis is known not to survive exposure to NaOH decontamination.

In 1986 and 1987 a few additional reports on mycobacteria and Crohn's disease appeared, but these were not research papers. Tytgat and Mulder (278) presented a review on the etiology of Crohn's disease and were the first to report, in other than abstract form, the isolation of M. paratuberculosis from a patient in the Netherlands. Data published in this review represented the first independent duplication of previous efforts and confirmation that M. paratuberculosis may be isolated from some cases of Crohn's disease. While all the active theories on the etiology of Crohn's disease were addressed, the authors considered that "a microbial aetiology, particularly mycobacterial, seems the most promising".

.... Haagsma et al presented data on cultivation of mycobacteria from patients with Crohn's disease (113) and the presence of M. paratuberculosis antibodies in Crohn's disease patients (114). They cultured 66 surgical specimens and isolated M. paratuberculosis from 1, M. fortuitum from 1, and acid-fast material from 2. Colonies of M. paratuberculosis emerged after 11 and 16 months incubation on Herrold's egg yolk and Ogawa media, respectively. These studies, however, had two major flaws: the processing protocol changed sometime during the study and control tissues were not cultured. Their isolates of M. paratuberculosis were found to be genetically identical to those isolated by Chiodini in the United States (56). These investigators have recently isolated an additional strain of M. paratuberculosis from a Crohn's disease patient, bringing the number of Crohn's disease-associated M. paratuberculosis isolates to 2 out of 88 specimens examined (Haagsma, J., personal communication, 1988).

....G. Gitnick, et al (98) described their efforts to isolate mycobacteria from resected Crohn's disease tissues and inoculation of animals with their organisms. These authors cultured tissues from 27 patients with Crohn's disease, 29 with ulcerative colitis, and 26 with other bowel diseases. Three strains of mycobacteria were isolated, of which two remain uncharacterized. One isolate from a patient with Crohn's disease was identified as M. chelonei, another was said to be similar to M. paratuberculosis, and the third isolate from a cancer patient remains uncharacterized. The two isolates from Crohn's disease required 3 and 12 months incubation, respectively. Acid-fast spherules were isolated from a few Crohn's disease patients as well as controls. The M. chelonei isolate was inoculated orally into newborn goats which subsequently developed a transient diarrhea. Three animals died 5-10 days post-inoculation. Intestinal lesions were limited to mild inflammation and colonic infiltration with polymorphonuclear cells. Animals receiving the uncharacterized M. paratuberculosis-like organism remained clinically and pathologically normal. Of interest is the apparent acute diarrheal disease produced by M. chelonei since this organism is generally associated with immunocompromised hosts or with traumatic wounds (31, 110, 111, 205, 263). An acute intestinal disorder produced by M. chelonei could have significant meaning to both the veterinary and medical professions. However, the authors did not adequately rule out other neonatal diseases of goats as a possible cause of the observed diarrhea and acute bowel inflammation. The latest data from this laboratory indicates that they have isolated mycobacteria from 3 out of 27 patients with Crohn's disease, 1 out of 31 ulcerative colitis patients, and 1 out of 27 controls. Two of the 3 isolates from Crohn's disease are M. paratuberculosis (one genetically confirmed); the other is the strain of M. chelonei reported above. The strains from controls and an ulcerative colitis patient are slow-growers, as of yet unidentified, but do not appear to be M. paratuberculosis (Gitnik, G., personal communication, 1988).

Lastly, at the American Gastroenterological Association meeting in May 1988, the abstracts related to mycobacteria and Crohn's disease were only genetic studies on some of the isolates. ....

Lastly, the Bovine Pathology Laboratory of the Lyon Veterinary School in France isolated a strain of M. paratuberculosis from a 45-year old woman with Crohn's disease. This isolate was identified by numerical taxonomic methods at the Laboratoire Central de Recherches Veterinaires. Drs. Descos and Perard of the Lyon-Suds Hospital and Lyon Veterinary School, respectively, have initiated a study to attempt isolation from fecal and biopsy specimens from approximately 50 patients with Crohn's disease.

It is now clear that a host of different mycobacteria can be isolated from Crohn's disease patients, as well as control populations, and that diseased tissue may be a suitable micro-environment for colonization of some of these species (Table 1 http://alan.kennedy.name/crohns/research/diagnos/chiodini.htm). Most of these organisms are environmental opportunists (Table 2 http://alan.kennedy.name/crohns/research/diagnos/chiodini.htm), although a few investigators have isolated the pathogenic M. paratuberculosis (Table 3 http://alan.kennedy.name/crohns/research/diagnos/chiodini.htm). Unfortunately, in all studies reported to date, different methods have been used (Table 4 http://alan.kennedy.name/crohns/research/diagnos/chiodini.htm) and as would be expected, many different results have been obtained. This is true not only for the cultural studies, but for immunological studies as well. .....

.... It is interesting to note that all investigators who have been successful in isolating M. paratuberculosis from Crohn's disease patients were trained originally in veterinary mycobacteriology and had years of experience dealing with this peculiar species.

.... If Crohn's disease patients are infected with low numbers of M. paratuberculosis or some other Mycobacterium species and super-infected with organisms similar to the LAM and LDC, such a situation could account for the data generated.

.... Despite the appropriate evaluation of cellular immunity in chronic conditions, such as Crohn's disease, most studies to date have examined humoral immunity and these have been quite limited. Except for a few scattered reports, immunologic studies related to mycobacteria and Crohn's disease have been conducted either in direct response to bacteriologic data (presented under Cultural Evidence), or involved the use of mycobacterial antigens in accessing general immunologic functions.

.... antigens in Crohn's disease by the ELISA technique. Patients with Crohn's disease had a statistically significant increase in antibody titer to a protoplasmic antigen of M. paratuberculosis as compared to controls. .... As a result of this cross-reactivity, a significant proportion of Crohn's disease patients' sera also reacted to M. kansasii antigens. These results have not been duplicated in any other laboratory.

Cho et al (57) recognized that these inconsistencies in seroreactivity did not necessarily contradict the on-going theories, particularly when organisms related to the MAI complex were involved.

Jiwa et al (130) described IgG serum antibodies to mycobacterial PPD's in Crohn's disease patients. These investigators examined seroreactivity to PPD's prepared from M. tuberculosis, M. kansasii, M. phlei, M. paratuberculosis, and M. smegmatis and found that Crohn's disease patients have elevated antibody titers to all species examined. ..... Such widespread reactivity to PPD, probably based on a ubiquitous cross-reactive antigen, is highly indicative of sensitization by environmental organisms gaining immune access through a defective mucosal barrier.

Treatment Data

Reports on the treatment of Crohn's disease with anti-mycobacterial agents are sporadic and generally not double-blinded nor well controlled with placebo treatment. Many are individual case reports ...... or involve few patients (216, 217, 274, 301). ....

Recently there has been a surge of interest in the treatment of Crohn's disease with anti-mycobacterial agents, which undoubtedly, has been precipitated by the suggestions of M. paratuberculosis as the etiologic agent of this disease. Of the studies recently conducted or in progress, most have used rifabutin (Adria Laboratories, Columbus; Farmatalia Carlo Erba, Milan), a rifampin derivative, either alone or in combination with other drugs. The reasons for the interest in rifabutin as the anti-mycobacterial drug of choice is due in part to its high in vitro activity against M. paratuberculosis (unpublished data) and its successful use in the treatment of paratuberculosis in subhuman primates (172). Additionally, the manufacturer of rifabutin has been very supportive of its use as a chemotherapeutic agent in Crohn's disease. Nevertheless, the data accumulated to date has been poorly organized and difficult to interpret.

....Although some minor effect may be occurring on drug-treated patients, the data presented is no very encouraging.

Thayer et al .... ) used rifabutin in combination with reptomycin in an open trial with 12 patients with Crohn's disease. Streptomycin (1-gram) was given intramuscularly 5-days a week for 2-4 months, and rifabutin was given orally at 300 mg/day for a minimum of 6-months or until drug withdrawal was elected. ..... All patients have reportedly improved clinically, commonly with prednisolone withdrawal, healing of fistularization, and marked improvement in their CDAI. Of the 12 patients, 8 (66%) have completely withdrawn from steroids, 2 of 2 (100%) have withdrawn from 6-mercaptopurine or other drugs, 7 of 7 (100%) no longer have rectal bleeding, 4 (33%) have endoscopic healing of lesions, and 1 of 2 (50%) had radiographic improvement. These improvements were generally not noted until after at least 4-months duration of treatment and were most prominent after 6-months. Some patients failed to respond until after 6-months of treatment, thereby illustrating the need for long-term therapy in this chronic disease. ...........

Discussion of Treatment Data

Although a mycobacterial etiology of Crohn's disease has been considered for well over 50 years, few studies have been conducted and little conclusive data are available on the effects, beneficial or not, of anti-mycobacterial chemotherapy. Often studies have been performed with little forethought and without supportive laboratory data. ......

..... Crohn's disease occurs with its highest incidence in the United States, the United Kingdom, and Scandinavia. It is less frequent in Central Europe and rarely is reported in Africa, Asia, and South America. The disease is seldom reported in underdeveloped or developing countries (35, 36). In the United States, the incidence is somewhere between 3.1 to 13.5 per 100,000 population, and is between 0.3 to 7.3 in other countries where the disease is reported (35, 36). Reports are conflicting, but the incidence of Crohn's disease in the United States and in other countries has been increasing, particularly in certain regions (35, 36, 91, 208, 262). Generally, the prevalence of disease appears to have stabilized in most countries. In contrast, tuberculosis (and leprosy) occur with highest frequency in those areas where Crohn's disease is rarely seen, and with low frequency where Crohn's disease is most frequent.

.... Some individuals have suggested that the apparent increase of Crohn's disease in the western world is related to the decreasing incidence of tuberculosis, because infection or immunization with one Mycobacterium species provides protection against infection with another species. ....

In Table 8 http://alan.kennedy.name/crohns/research/diagnos/chiodini.htm,

......There is a known familial association of Crohn's disease (84, 144, 157, 170, 259, 290) which suggests a genetically-linked increased susceptibility to the disease, or alternatively, a common exposure to an etiologic agent. There is a low incidence of Crohn's disease in married adults, but these rare occurrences have yet to be explained.... There is a 30 times greater rate of Crohn's disease in siblings and 13 times greater incidence in first degree relatives (84, 85). Such a familial association, the occurrence of Crohn's disease in siblings and mono- and di-zygotic twins (including those living apart since early childhood), and the rarity of Crohn's disease in half-siblings ...), indicates a genetic susceptibility or predisposition occurring as a recessive trait.

.... A high proportion of Crohn's disease patients have auto-antibodies against gastrointestinal tissue and other self antigens (8, 260). Although these anti-colon antibodies may arise due to cross-reactivity with E. coli 014 K1 antigens (260), the documentation is rather weak. Circulating immune complexes are also observed in Crohn's disease (67, 99, 153). ,,,,, Both the presence of low level auto-antibodies and circulating immune complexes are common manifestations of chronic disease in general.

...... Non-steroidal anti-inflammatory agents are known to activate quiescent Crohn's disease (140) and induce intestinal inflammation in other types of patients as well (24). Therefore, their use is contraindicated.

In summary, chemotherapeutic schemes offer several similarities between the mycobacterioses and Crohn's disease: i. anti-mycobacterial agents appear effective in only a portion of Crohn's disease patients as they are in intestinal mycobacterioses; ii. corticosteroids offer clinical improvement in Crohn's disease,... and other mycobacterioses; and iii. non-steroidal anti-inflammatory agents activate quiescent Crohn's disease and tuberculosis. Despite these similarities, the efficacy and appropriateness of antimycobacterial chemotherapy in Crohn's disease remains to be evaluated.

...... It is highly unlikely that mycobacteria cause all cases of Crohn's disease, but available data suggest strongly that they do cause some cases. The mycobacterial etiology theory of Crohn's disease remains alive.