MYCOPLASMA-LIKE ORGANISMS ANDOPHTHALMIC DISEASE*

BY Emil Wirostko, MD (BY INVITATION), Lewis A. Johnson, MD

(BY INVITATION), Barbara M. Wirostko, MD (BY INVITATION), AND

R. Linsy Farns, MD

CLASSIC BACTERIOLOGY: A HISTORICAL PERSPECTIVE

DURING THE 19TH CENTURY GREAT STRIDES WERE MADE IN THE UNDER-

standing of human pathogenic bacteria.1 Culture techniques were devel-oped, microscopic morphologic characteristics were elucidated, and induc-tion of animal disease was accomplished.' Much of this was the result ofpioneering work of Pasteur and Koch.' For decades the greatest reliancehas been placed on the culture techniques in the detection and characteriza-tion of bacteria.2 Bacteriologists now recognize that most bacteria in naturedefy cultivation.3 Molecular biologic techniques are now widely used tosupplement our knowledge.3'4 Various human bacterial pathogens havebeen difficult to study and characterize because they have defied cultureand transmission to experimental animals. A well-known example of this isthe leprosy bacillus, which is readily detectable by light microscopy of thedlseased tissues.5 Another example is the Whipple bacillus, which is onlydetectable with certainty in the diseased tissues by transmission electronmicroscopy.6 It is important to note that Amsler7 reported in 1955 thatsterile uveitis intraocular fluids from many patients demonstrated bacteria-like structures that resisted cultivation. In 1974 we reported similar observa-tions.8 We also reported that intraocular fluids containing these structuresproduced chronic uveitis in experimental animals.8 Electron microscopy ofthese sterile fluids is important in the understanding of the uveitis bacte-ria.9-11

Bacteria can be classified as either "Firmicutes, Gracillicutes, or Mol-licutes" on the basis of their ultrastructural features.12 Firmicutes are gram-positive owing to their thick cell wall, and this is readily apparent.13 Gra-

'From the Columbia-Presbyterian Medical Center and Edward S. Harkness Eye Institute, andthe Department of Pathology, New York. Supported by the Milton and Grace Johnson FamilyTrust.

Wirostko et al

cillicutes are gram-negative, owing to the absence of thick glycopeptide cellwall material, the presence of an undulating, delicate cell wall, and a distinctperiplasm separating the cell wall from the plasma membrane.'3 Mollicuteslack a cell wall.'4 They are enclosed only by a plasma membrane.'4 Mol-licutes are further characterized by extreme fastidiousness, markedly pleo-morphic nature, and a weak affinity for the usual biologic stains.14,15 More-over, mollicutes pass bacteria-retaining filters.'4 Mollicutes can be easilyoverlooked or mistaken for viruses.14 In 1967 Japanese investigators re-ported that chronic plant disease long thought to be viral in nature is causedby intracellular mollicutes.'6 Cells parasitized by these mollicutes displaydysfunction, destruction, and/or proliferation.'6"17 Despite over 25 years ofeffort these mollicutes have defied cultivation.'6 The bacterial nature andpathogenicity of these mollicutes have been confirmed by transmissionstudies and their response to antibiotics.'6 The inability to cultivate thesemollicutes precluded their speciation for many years.16 Consequently, theywere given the eponym "mycoplasma-like organisms," or simply MLO.16With use of molecular biologic techniques the phylogeny of these mollicuteshas been elucidated and several subtypes have been identified.'8 Thesemollicutes are somewhat closely related phylogenetically to the Acholeplas-ma and only distantly to the other extracellular cultivable mollicutes (Fig

ANEROPLASMA SPI ROPLASMA

UREAPLASMA

MYCOPLASMA

MLO

ACHOLEPLASMA

FIGURE 1

Phylogeny of class mollicutes (based on 16S rRNA data).

CLASSFIRMACUTES

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Mycoplasma-Like Organisms87

MYCOPLASMA-LIKE ORGANISMS IN HUMAN OPHTHALMIC DISEASE

Inflamed intraocular fluids from human chronic uveitis may display leuko-cytes with abnormal intracellular bodies indistinguishable from plant MLO(Fig 2).9- 1,20-23 Contemporary culture techniques of these fluids fail toexhibit propagating bacteria or viruses.9- 1,20-23 Molecular biologic tech-niques show that these human MLO are phylogenetically closely related to,yet distinct from, plant MLO (B. C. Kirkpatrick, personal communication,1993). Inoculation of these MLO into the eyelids of young mice producedchronic ophthalmic inflammatory disease, including uveitis, after a 3-monthlatent period (Figs 3 and 4).24-26 MLO could be demonstrated in theinflamed ocular tissues.9-1124-26 With the onset of the eye disease the

FIGURE 2Mycoplasma-like organisms (MLO) parasitized human chronic uveitis leukocyte. Cytoplasm ofa vitreous monocyte is filled with MLO (arrows) (uranyl acetate-lead citrate, x 22,780) (from

Wirostko et a122).

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88 Wirostko et al

: ~~ ~ ~ ~ ~ ~~~~- k.

FIGURE 3MLO-induced murine chronic uveitis. Choroid and vitreous of mouse dying 7 months afteruveitis MLO inoculation into eyelids display infiltrate of small mononuclear leukocytes. Macro-phages are present beneath detached retina. MLO parasitized leukocytes were abundant in

lesion on electron microscopy (hematoxylin-eosin, x 106) (from Wirostko et al25).

animals exhibited accelerated mortality.25 Death was due to randomlydistributed chronic inflammatory disease in vital organs, such as the heart,27lungs,28 liver,29 and gastrointestinal tract.30 MLO were present in theophthalmic and visceral disease.24-30

Pathobiologic features of MLO disease are of interest. The experimentaldisease is initially a microvasculitis (Fig 5).27-31 Parasitization of leukocytesand endothelial cells appears to be important in the induction of this lesion(Fig 6).31 With disease progression one sees extensive lymphocytic infil-trates, tissue lysis, and occasional granuloma formation (Fig 7).27-30 It has

Mycoplasma-Like Organisms 89

40 4~ ~~~~FGUE

MLO-induced murine orbital lyiphoid pseudotumor. Extensive orbital ononuclear. leuko-cytic infiltrate in mouse dying 9 months after MLO inoculation into eyelids was accompaniedby exophthalmus. MLO parasitized leukocytes were readily found in infiltrate by electron mi-

croscopy (lbematoxylin-eosin, x 92) (fromn Wirostko et a124).

been suggested that MLO-induced leukocyte dysfunctionminay be responsi-ble for disease progression and the appearance of autoimmune phenom-ena.9-1127-30 Foulds32 has recently reported an innovative technique forretinal biopsy. He has reported finding MLO parasitized retinal pigmentepithelial cells, along with MLO parasitized leukocytes in the ocular dis-ease.32 The observations of Foulds, as well as our observations, suggest thatMLO can parasitize a wide range of cells.9-11,31,32There are many reasons to believe that MLO could also be involved in

human idiopathic systemic inflammatory disease.2t-23273t) Many of the

Wirostko et al

FIGURE 5MLO-induced microvasculitis. Small vessel in lethal carditis induced by uveitis MLO inocula-tion into mouse eyelids displays mononuclear leukocytic infiltrate. Endothelial cells are swollenand basement membrane is disorganized. Many leukocytes exhibited MLO parasitization by

electron microscopy (uranyl acetate-lead citrate, x 5176) (from Johnson et a31).

patients with MLO uveitis that we have studied also had idiopathic inflam-matory bowel disease.20,'21 Such bowel disease is frequently associated withchronic uveitis.20,21 Foulds has also reported finding MLO in the eyedisease of patients with inflammatory bowel disease.33 In the mouse MLOproduces a multisystemic disease including eye and gut involvement.24-26,30We and others are investigating the possibility that MLO are involved inhuman inflammatory bowel disease.

In our experience, MLO are a common cause of human chronic ophthal-mic disease.20-23 Finding MLO as a cause of human eye disease suggests

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Mycoplasma-Like Organisms

aS

FIGURE 6MLO parasitized endothelial cells. Two endothelial cells in cardiac microvasculitis induced byuveitis MLO inoculation into mouse eyelids display innumerable intracytoplasmic MLO. Onlyremnant of nucleus of lower cell is detectable (arrow) (uranyl acetate-lead citrate, x 13,145)

(from Johnson et aPI).

that antibiotics could be a useful addition to therapy.2t-23 In our experience,tetracyclines and erythromycin have a limited effectiveness in human MLOdisease. Long-term rifampin therapy in uncontrolled human studies appearsto be beneficial.8 It is of interest that in experimental MLO disease, rifam-pin significantly decreased the induced mortality when compared with thecontrols (Fig 8).8 Active incorporation of rifampin into leukocytes may be afactor in rifampin's efficacy.2030 It is of interest that rifampin has recentlybeen found to be highly effective against plant MLO disease (B. C. Kir-patrick, personal communication, 1993).

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FIGUIRE 7Extensive MLO disease. Lung of a mouse dying several months after eyelid inoculation withuveitis MLO displays chronic interstitial pneumonitis, alveolar septal lysis, and granulomaformation. MLO parasitized leukocytes were readily apparent in infiltrate by electron micros-

copy (hematoxylin-eosin, x 212) (from Wirostko28).

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Mycoplasma-Like Organisms

100

75 NN 3K

(0 3

25

0 _3 6 9

MONTHSFIGURE 8

Significant reduction of MLO-induced murine mortality by rifamplin. Pyramids indicate 60MLO-inoculated, saline-treated mice; squares indicate 60 MLO-inoculated, rifamplin-treatedmice; circles indicate 60 saline-inoculated, rifampin-treated control mice. Mortality differencesof pyramid group versus both square and circle groups at 9 months = < 0.05. All animals weredosed once daily per OS. Also note delayed onset of MLO-induced mortality (from Wirostko

and Johnon8).

REFERENCES

1. Joklik WK, Wheat RW, Willett HP: Zinsser Microbiology. Norwalk, Appleton & Lange,1992, pp 3-7.

2. Wheat RW: Zinsser Microbiology. Norwalk, Appleton & Lange, 1992, pp 8-17.3. Olsen GJ, Lane DJ, Giovannoni SJ, et al: Microbial ecology and evolution: A ribosomal

RNA approach. Annu Rev Microbiol 1986; 40:337-365.4. Razin S: The Mycoplasmas, vol 5. San Diego, Academic Press, 1989, pp 33-69.5. Willett HP: Zinsser Microbiology. Norwalk, Appleton & Lange, 1988, pp 441-445.6. Johnson LA, Diamond I: Cerebral Whipple's disease: Diagnosis by brain biopsy. AmJ Clin

Pathol 1980; 74:486-490.7. Amsler M: L'humeur Aqueuse et ses Fonctions. Paris, Masson, 1955, pp 218-243.8. Wirostko E, Johnson L: The induction of mouse uveitis by human idiopathic uveitis

aqueous humor and treatment by certain antituberculous drugs. Trans NY Acad Sci (SeriesII) 1974; 36:693.

9. Johnson L, Wirostko E, Wirostko B: Chronic idiopathic vitritis: Cytopathogenicity ofunusual bacteria for vitreous polymorphonuclear leukocytes. J Submicrosc Cytol Pathol1987; 19:161-166.

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10. Wirostko E, Johnson L, Wirostko W: Chronic leukocytoclastic bacterial vitritis: A lympho-cyvte transmission electron microscopic study. J Stubmicrosc Cytol Pathol 1987; 19:651-656.

11. Chroniic initracellular leukocytoclastic bacterial vitritis: A transmission electronmicroscopic study of the monocytes. J Subbmicrosc Cytol Pathol 1988; 20:463-470.

12. Tully JC: The Mycoplasnmas, vol 5. San Diego, Academic Press, 1989, pp 1-31.13. Wheat RW: Zin.sser Microbiology. Norwalk, Appleton & Lanige, 1992, pp 18-30.14. Boatman ES: The Mycoplas1ma.s, vol 1. Newv York, Academic Press, 1979, pp 63-102.15. Rodwell AW, Mitchell A: The AMycoplasmas, vol 1. New York, Academic Press, 1979, pp

103-109.16. McCoy RE, Catudwell A, Chang CJ, et al: The AMycoplasmnas, vol 5, San Diego, Academic

Press, 1989, pp 545-563.17. Saglio PMHI, Whitcomb RE: The Alycopla.smas, vol 3. New York, Academic Press, 1979, pp

1-36.18. Kuske CR, Kirkpatrick BC: Identificationi anid characterizationi of plasmids from the

Western Asters Yellows mycoplasmalike organiism. J Bacteriol 1990; 172:1628-1633.19. Kirkpatrick BC, Sears B, Seemuller E, et al: Internationial Organization for Mycoplasnlol-

ogy Letters, vol 2. Ames, Iowa, IOM, 1992, pp xiii.20. Johlnson LA, Wirostko E, Wirostko WJ: Crohln's disease uiveitis: Parasitization of vitreous

leuikocytes by mollictute-like organisms. Am J Clin Patlhol 1989; 91:259-264.21. Wirostko E, Johnson L, Wirostko B: Ulcerative colitis associated uveitis: Parasitization of

intraocular leucocytes by molliciute-like organisms. J Subhnicrosc Cytol Pathol 1990; 22:231-239.

22. Wirostko E, Johnson L, Wirostko W: Juvenile rheumatoid arthritis inflammatory evedisease: Parasitization of ocular leukocytes by mollicute-like organisms. J Rhleiumrlatol 1989;16: 1446-1453.

23. Wirostko E, Johnson L, Wirostko B: Sarcoidosis associated uveitis: Parasitization of vit-reous leucocytes by mollicute-like organisms. Acta Ophlthalmol 1989; 67:415-424.

24. Mouse exophthalmic chronic orbital inflammatory disease: Indluction by hullmanleucocyte intracelltular mollicutes. Virchows Arch [A] 1988; 413:349-355.

25. Wirostko E, Johnson LA, Wirostko BM: Transmission of chronic idiopathic vitritis in miceby inoculation of human vitreouis containing leucocyte phagolysosomal bacteria-like bod-ies. Lancet 1986; 8505:481-483.

26. Wirostko E, Johnson L, Wirostko B: Postinflammatorv cataracts in the mouise: Inductioni byhuman mycoplasma-like organismns. Br J Ophthalmol 1991; 75:671-674.

27. Johnsoni L, Wirostko E, Wirostko W: Mouse lethal cardiovascular disease induction byhuman leucocyte intracellular mollicutes. Br J Exp Pathlol 1988; 69:265-279.

28. Wirostko E, Johnson LA, Wirostko WJ: Mouse interstitial lung disease and pletiritisinduction by humain mollicuite-like organisms. Br J Exp Pathol 1988; 69:891-902.

29. Johlnson L, W'irostko E, Wirostko W: Primary biliary cirrhosis in the mouse: Induction byhuman mycoplasma-like organisms. Int J Exp Pathol 1990; 71:701-712.

30. Johnson L, Wirostko E, Wirostko W, Cummings W: Experimiiental murinie chronic inflam-matory bowel disease: Induction by human uiveitis mvcoplasma-like organisms. Pathol ResPrac 1993. In press.

31. Johnson L, Wirostko E, Wirostko W, et al: Mycoplasma-like organism induced murinecardiac microvasculopathy: A tranismissioni electron microscopic study. Pathol Res Prac1993; 189:448-452.

32. Fouilds WS: The uses and limitations of intraocular biopsy. Eye 1992; 6:11-27.33. Lee WR: Mollicutes in the retinal pigment epithelium. Presented at the Proceedings of tcle

European Ophlthlalmlic Pathlology Society. Juniie 3-6, 1990; Coimbra, Portugal.

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DISCUSSION

DR THOMAS J. LIESEGANG. Almost 100 years ago pleuropneumonia organisms wereisolated from cattle, and subsequently the same organisms were isolated from otheranimals and from humans. They were initially labeled Mycoplasnw but later the term"mollicutes" were established for this class of organisms and Mycoplasnm is a genuswithin this class. They are prokaryotic (ie, unicellular), with no cell wall and nodistinct nucleus. They are the smallest prokaryote capable of self-replication. Theyare distinct from unstable bacterial L forms.

Extracellular mollicutes are a frequent cause of human atypical pneumonia and ofopportunistic infections in the mouth and urogenital system. These extracellularmycoplasmas are fastidious but can be grown in the laboratory, are lipid-rich, andcontain a variety of cytotoxic substances, including nucleases.

Intracellular mollicutes have been observed by transmission electron microscopysince the 1970s and are an established cause of various plant diseases (The Mycoplas-mas, vol 5. San Diego, Academic Press, 1989, pp 545-640). They have never beencultured, so their taxonomy has been uncertain, although recent investigations placethem in the class phytoplasmas.The work of these authors spands 20 years in at least 20 peer-reviewed articles. In

1974 they noted small organisms in human uveitis fluid by tranmission electronmicroscopy (TEM). They expanded their studies culminating in detailed reports inthe mid-1980s of a "mycoplasma-like organism" (MLO) within vitreous lympho-cytes. By TEM only, they have identified thick-walled coccal bodies and electron-dense tubulospherical bodies or filaments within a small proportion (ie, 2% to 10%)of neutrophils, lymphocytes, or monocytes in some diseased tissues. They hypothe-size that the smaller elementary bodies evolve to the larger bodies, but they have notprovided proof. Material from four patients with idiopathic vitritis was injected intothe eyelids of mice, resulting in ocular disease consisting of uveitis, cataract, andorbital inflammation, as well as liver, lung, and lethal cardiac disease. The MLO wereidentified in a small proportion of leukocytes in the diseased tissues of these mice.Appropriate control studies were performed simultaneously, and the scientific proto-col appears valid. Mouse-to-mouse passage has been performed.The authors have further extended TEM to other disease processes and have

subsequently identified MLO in the vitreous of patients with the uveitis associatedwith Crohn's disease, sarcoid, and juvenile rheumatoid arthritis, as well as in theorbital tissues of patients with nonspecific chronic orbital inflammatory disease.

These investigators suggest that MLOs can parasitize lymphocytes, monocytes,and neutrophils in humans and subsequently replace the cytoplasm, destroy theorganelles, and alter the nucleus, resulting in either cell proliferation, cell destruc-tion, or cell dysfunction. The initial resulting pathologic lesion has been charac-terized as a vasculitis with subsequent tissue lysis, lymphoid infiltration, and granu-loma formation.These studies present compelling data, but let me raise some cautions, some

suggestions, and some parallels from other diseases. Although MLOs undoubtedlyinvade and multiply in plant cells, their invasion into animal or human cells has not

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been readily embraced by the community of microbiologists or pathologists. Mouse-to-mouse transfer has been demonstrated. The sequence of change from elementarybody to these MLO organisms has not been demonstrated. There is even thepossibility that these TEM observations are unusual cellular organelles, rather than amicroorganism, or that the structures are a consequence, rather than a cause, of theobserved inflammation.

These organisms are difficult to detect in the absence of a significant number ofleukocytes, and other techniques to detect them need to be pursued. Cultureconditions that simulate the nutritional milieu of the host need to be explored,although 20 years of effort have been unsuccessful to date. The absence of anorganism free of the host cell hampers the development of serodiagnostic tests. Theinvestigators of plant MLOs have described a bioassay technique using insects (AnnMicrobiol [Paris] 1984; 135A:255-261), an assay of MLO nucleases following sonicrupture of parasitized plant cells (J Bacteriol 1985; 164:811-815), and a polymerasechain reaction for detecting MLO (Biochem Biophys Res Coinmun 1992; 186:1503-1509). It is obvious that other medical centers with access to TEM need to confirmthe findings of these authors, especially in view of the multiple disease entitiesreported to be associated with MLO.A number of independent observers, however, are beginning to report "MLO

sightings." Foulds (Eye 1992; 6:11-27) has detected similar MLOs in the retinalpigment epithelial cells of a patient with intraocular inflammation and reported aresponse to rifampin. Mycoplasrna penetrans and Mycoplasmafermlentans are bothseen in patients with AIDS and are the first true mycoplasmas shown to be capableof penetrating human cells (Int J Syst Bacteriol 1992; 42:357-364 and FEMSMicrobiol Let 1992; 100:423-432). Both are able to be cultured in the laboratory.Mycoplasma fermentans is probably causative in AIDS-associated nephropathy.MLOs have been detected by other investigators in the conjunctival epithelial cellsin association with swine conjunctivitis (J Am Vet Med Assoc 1991; 198:450-452), thesmall bowel of children with chronic diarrhea (Scanning Microsc 1991; 5: 1037-1062),in an insect disease with transmissible sterility (Pathogenic Mycoplasmas. Amster-dam, Elsevier, 1972, pp 227-250), and in an infection in shrimp (J Invertebr Pathol1991; 57:362-370).

Rifampin is a broad-spectrum antibiotic that is active against a wide variety ofintracellular pathogens and does concentrate in leukocytes. It has been utilized forprolonged administration and has been used empirically for inflammatory boweldisease and in selected cases of mouse and human uveitis. Other investigators needto confirm and explore the leads presented by these investigators.

DR RICHARD DARRELL. This is an excellent and stimulating paper. You mentionedmolecular probes. How do the MLO organisms compare with other mycoplasma by16S RNA sequences?

DR LEONARD APr. On listening to this paper I couldn't help but think back toexperiences I had as a resident at Wills Eye Hospital many years ago while rotatingthrough the Uveitis service headed by Dr Leopold. At that time we knew nothing

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about mycoplasma-like organisms (MLO). I vividly recall several patients, who, toour surprise, responded favorably to an intensive course of Aureomycin (chlor-tetracycline)-a fairly new broad-spectrum antibiotic. We had no specific reason togive the antibiotic except we reasoned that since the patients were not improvingwith the usual nonspecific therapy, perhaps we were dealing with an unidentifiedinfectious agent. Now that I am familiar with what MLO can do in the eye, I wonderwhether we were treating MLO uveitis.The tetracyclines, and more recently, rifampin, have been found effective, at least

to some extent, against MLO. Hence, in these days it may be wise for clinicians toconsider using intensive antibiotic therapy, including anti-MLO drugs, empirically totreat uveitis patients in whom the cause is unclear or who are not responding to othertherapy.

Recently, a moderately new monocyclic beta-lactam antibiotic, aztreonam, hasshown some promise against MLO. I ask Dr Wirotsko whether she and her associ-ates have used this drug in their studies.

I also ask Dr Wirotsko about her current thoughts on MLO as causative agents inhuman cataracts. In a paper her group published in 1991 (BrJ Ophthalmol 75:671-674) titled "Postinflammatory cataracts in the mouse: Induction by human mycoplas-ma-like organisms," they suggested that MLO could cause human cataracts.

I enjoyed hearing this paper.

DR BARBARA WIROSTKO. Thank you for your comments. Thank you, Dr Liesegang,for a very interesting discussion and review of our work. As Dr Liesegang noted, bothextracellular and intracellular mycoplasm forms exist. Addressing Dr Darrell's ques-tion, we are presently using molecular biologic techniques to determine the genesequences of the MLO we find in human ophthalmic disease. In collaboration withDr Bruce Kirkpatrick, a plant pathologist at the University of California, Davis, weare finding that human MO have a 92% homology with plant MLO. Moreover, the16S rRNA oligonucleotide sequences we are finding in human MLO have only aboutat 40% homology with the extracellular cultivable mycoplasma. Therefore, themolecular biologic assays confirm the presence in human ophthalmic disease of aprokaryotic organism with morphologic and pathobiologic properties extremelyclosely related to plant MLO. We wish to emphasize, however, that an investigatorunfamiliar with the ultrastructural features of bacteria and normal cells couldoverlook the presence ofMLO in the eukaryotic host cells. Dr Thomas Chen, a plantpathologist at Rutgers University, has produced four monoclonal antibodies to highlypurified plant MLO extracts. None of these antibodies reacted with our humanMLO samples. In future investigations we hope to produce sensitive highly specificmonoclonal antibodies to human MLO. Such substances could have diagnosticutility. The possibility that human MLO may be cultivable needs to be investigatedfurther. Dr Shih Lo at the Armed Forces Institute of Pathology, Washington, DC,has been successful in cultivating on cell-free media a host cell parasitizing myco-plasma that is frequently found in AIDS patients. We have not tried his media, butwe plan to do this soon. However, the close genetic homology of plant and humanMLO suggests that cultivation of the human organisms in cell-free media may not be

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possible. Our future attempts at in vitro cultivation of buman MLO will be focusedprimarily on long-term stable leukocyte, endothelial cell, and retinal epithelial cellcultures.

As Dr Liesegang noted, many patients with uveitis also have inflammatory boweldisease. Gastroenterologists are aware that inflammatory bowel disease often re-sponds favorably to long term rifampin administration. It is interesting that ProfessorWallace Foulds, University of Glasgow, has reported that rifampin dramaticallyimproved the vision of a Crohn's disease patient with uveitis in whom he found MLOparasitized retinal pigment epithelial cells. In our experience rifampin must be givenfor a prolonged period for best results.Dr Apt made an interesting point about tetracycline therapy and mycoplasmal

disease. Plant MLO disease does indeed respond to long-term tetracycline adminis-tration. We have not found either tetracycline or erythromycin to be effective inhuman MLO ophthalmic disease. We have not tried either of these two antibiotics inthe mouse model MLO disease. However, it is interesting that Barbara Sears, a plantpathologist at Michigan State University, has recently found that rifampin is just aseffective as tetracycline against plant MLO. Dr Apt asked if we have tried the newantibiotic Aztreonam against human MLO. We have no experience with this antibi-otic in human MLO disease, but it should be tried.The possibility that human MLO could be involved in cataract induction, partic-

ularly with postinflammatory variety, was raised. Laboratory mice receiving eyelidinoculations of human uveitis MLO develop a high incidence of inflammatorycataracts. Electron microscopy of lens tissues is technically difficult in our experi-ence. However, fixation of the lens in a mixture of 2% formaldehyde/2% glu-taraldehyde provides material that can be studied by both light and electron micros-copy. Excision of foci rich in leukocytes from the paraffin block of the complicatedcataract will provide samples that can be reimbedded in plastic for electron micros-copy. Our report in the Bnitish Joutrnal ofOphthalmology 1991; 75:671-674, providesdetails on this methodology, and gives details of the MLO within parasitized intra-lenticular leukocytes. Human postinflammatory cataracts need to be studied forMLO using the transmission electron microscope and molecular biologic tech-niques. IfMLO are found in human postinflammatory cataracfs, such material mightbe used to induce cataracts in laboratory animals.

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