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Tuberculous Uveitis: Distribution of Mycobacterium tuberculosis in the Retinal Pigment Epithelium FREE

Narsing A. Rao, MD; Sindhu Saraswathy, PhD; Ronald E. Smith, MD
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Section Editor: W. Richard Green, MD

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Arch Ophthalmol. 2006;124(12):1777-1779. doi:10.1001/archopht.124.12.1777.
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The diagnosis of intraocular tuberculosis has been a challenge until the introduction of polymerase chain reaction (PCR) to detect the mycobacterial-specific DNA sequence in the intraocular fluids. Such investigations have confirmed diverse manifestations of intraocular tuberculosis including retinal vasculitis and serpiginouslike choroiditis.1,2 The latter may manifest with multifocal lesions involving the inner choroid, which coalesce to form a geographic pattern simulating serpiginous choroiditis. Although, clinically, the choroiditis appears to involve primarily the inner choroid and retinal pigment epithelium (RPE), the presence of mycobacterium at these anatomic sites has not been documented. Herein, we describe a case of panuveitis that was clinically of unknown cause but for which histopathologic examination of the globe disclosed selective distribution of acid-fast organisms in the RPE. These were confirmed to be Mycobacterium tuberculosis by microdissection of the RPE, followed by real-time PCR.

The enucleated left eye of a 48-year-old woman was submitted to the Doheny Eye Pathology Laboratory with a brief clinical history, revealing a progressively worsening intraocular inflammation in the left eye despite treatment with topical and systemic corticosteroids. The eye showed keratic precipitates, vitreous “snowballs,” and haze and retinal exudates. Laboratory studies included normal chest radiography results, purified protein derivative induration of 4 mm, and negative VDRL and Toxoplasma antibody test results while she was taking systemic corticosteroids. The clinical diagnosis was panuveitis.

Histologic examination of the globe revealed granulomatous inflammation of the uvea with necrosis of the retina and RPE (Figure 1A). Ziehl-Neelsen acid-fast stain revealed several acid-fast bacteria localized in the necrotic RPE cells, which stained positive for cytokeratin and neuronal-specific enolase and negative for CD68 (Figure 1B).

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Figure 1.

Histologic examination of the globe. A, Necrotic retinal pigment epithelium (RPE) with granulomatous inflammation involving the choroid (hematoxylin-eosin). The insert reveals several acid-fast bacteria in the RPE (Ziehl-Neelsen). B-D, The immunohistochemistry of the necrotic RPE cells. The cells are negative for CD68 (B) and positive for neuronal-specific enolase (C) and cytokeratin (D).

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The DNA was extracted from the RPE by microdissection of 5-μm-thick, paraffin-embedded sections of the globe. Microdissected RPE from a melanoma specimen was used as a control. The quantitative real-time PCR was performed using the forward primer IS6 (5′ AGGCGAACCCTGCCCAG-3′) and reverse primer IS7 (5′ GATCGCTGATCCGGCCA-3′), which amplified a 122–base pair fragment of the IS6110 multicopy element (GenBank accession No. X52471).3 The standard DNA used was 10 ng to 10 pg of M tuberculosis genomic DNA (Colorado State University). The amount of the test sample product was interpolated from the standard curve of cycle threshold values generated from known starting concentrations of standard M tuberculosis DNA. An average of 3 different runs revealed about 1.7 × 106 bacterial genomes in the RPE layer. The controls were negative for DNA amplification (Figure 2).

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Figure 2.

Amplification of Mycobacterium tuberculosis IS6110 multicopy element (GenBank accession No. X52471) insertion sequence with SYBR Green I. Fluorescent vs cycle threshold values from tuberculosis (TB) DNA from standard strain (genomic DNA from M tuberculosisH37RV) (A) and from test samples (C). Melting curve analysis of M tuberculosis amplification products from standard DNA (B) and test samples (D) indicating the specificity of the reaction product. Standard curve generated by analysis of known amounts of standard M tuberculosis genomic DNA with the Bio-Rad iCycler optical detecting system (Bio-Rad Laboratories, Hercules, Calif) (E). Purified DNA from the paraffin sections of test, control, and standard DNA were assayed in parallel. PCR indicates polymerase chain reaction; CF, curve fit; RFU, relative fluorescence units; dT, derivative of temperature; and Exp, experimental.

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The present clinicopathologic study combined with microdissection and real-time PCR analysis clearly revealed distribution of the mycobacteria in the RPE even though the retina and uvea were involved with the inflammatory process. Such findings suggest preferential localization of M tuberculosis in the RPE in eyes with panuveitis or related intraocular inflammation, including multifocal or serpiginouslike choroiditis resulting from tuberculosis.

In the pathogenesis of pulmonary tuberculosis, the mycobacteria are taken up by the alveolar macrophages that express complement and toll-like receptors.4 The bacteria are usually destroyed in the phagosomes when they fuse with lysosomes, exposing the bacteria to acid PH, reactive nitrogen species, and lysosomal enzymes.4 However, the mycobacteria can inhibit the fusion, thus avoiding the microbicidal activity, and may then thrive in the phagosome by avoiding immune surveillance by the T lymphocytes. Since RPE shares several functions with the macrophages, including phagocytosis of bacteria and expression of toll-like and complement receptors, the bacteria noted in the RPE may represent the phagocytosed bacteria.5 Moreover, the presence of numerous organisms in the RPE may suggest that the organisms thrive in the RPE by preventing phagolysosome fusion. Furthermore, the present report suggests that recurrences in tuberculous choroiditis could result from reactivation of sequestered organisms in the RPE. Prevention of such recurrences and elimination of the sequestered organisms require a longer course of treatment with systemic antimycobacterial agents, preferably at least 6 to 9 months.

Correspondence: Dr Rao, Doheny Eye Institute, 1450 San Pablo St, DVRC-211, Los Angeles, CA 90033 (nrao@usc.edu).

Financial Disclosure: None reported.

Funding/Support: This study was supported in part by a grant from Research to Prevent Blindness, New York, NY, and core grant EY03040 from the National Eye Institute.

Biswas  JShome  D Choroidal tubercles in disseminated tuberculosis diagnosed by polymerase chain reaction of aqueous humor: a case report and review of the literature. Ocul Immunol Inflamm 2002;10293- 298
PubMed
Gupta  VGupta  AArora  SBambery  PDogra  MRAgarwal  A Presumed tubercular serpiginouslike choroiditis: clinical presentations and management. Ophthalmology 2003;1101744- 1749
PubMed
Broccolo  FScarpellini  PLocatelli  G  et al.  Rapid diagnosis of mycobacterial infections and quantitation of Mycobacterium tuberculosis load by two real-time calibrated PCR assays. J Clin Microbiol 2003;414565- 4572
PubMed
Smith  I Mycobacterium tuberculosis pathogenesis and molecular determinants of virulence. Clin Microbiol Rev 2003;16463- 496
PubMed
Kumar  MVNagineni  CNChin  MSHooks  JJDetrick  B Innate immunity in the retina: toll-like receptor (TLR) signaling in human retinal pigment epithelial cells. J Neuroimmunol 2004;1537- 15
PubMed

Figures

Place holder to copy figure label and caption
Figure 1.

Histologic examination of the globe. A, Necrotic retinal pigment epithelium (RPE) with granulomatous inflammation involving the choroid (hematoxylin-eosin). The insert reveals several acid-fast bacteria in the RPE (Ziehl-Neelsen). B-D, The immunohistochemistry of the necrotic RPE cells. The cells are negative for CD68 (B) and positive for neuronal-specific enolase (C) and cytokeratin (D).

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Place holder to copy figure label and caption
Figure 2.

Amplification of Mycobacterium tuberculosis IS6110 multicopy element (GenBank accession No. X52471) insertion sequence with SYBR Green I. Fluorescent vs cycle threshold values from tuberculosis (TB) DNA from standard strain (genomic DNA from M tuberculosisH37RV) (A) and from test samples (C). Melting curve analysis of M tuberculosis amplification products from standard DNA (B) and test samples (D) indicating the specificity of the reaction product. Standard curve generated by analysis of known amounts of standard M tuberculosis genomic DNA with the Bio-Rad iCycler optical detecting system (Bio-Rad Laboratories, Hercules, Calif) (E). Purified DNA from the paraffin sections of test, control, and standard DNA were assayed in parallel. PCR indicates polymerase chain reaction; CF, curve fit; RFU, relative fluorescence units; dT, derivative of temperature; and Exp, experimental.

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Tables

References

Biswas  JShome  D Choroidal tubercles in disseminated tuberculosis diagnosed by polymerase chain reaction of aqueous humor: a case report and review of the literature. Ocul Immunol Inflamm 2002;10293- 298
PubMed
Gupta  VGupta  AArora  SBambery  PDogra  MRAgarwal  A Presumed tubercular serpiginouslike choroiditis: clinical presentations and management. Ophthalmology 2003;1101744- 1749
PubMed
Broccolo  FScarpellini  PLocatelli  G  et al.  Rapid diagnosis of mycobacterial infections and quantitation of Mycobacterium tuberculosis load by two real-time calibrated PCR assays. J Clin Microbiol 2003;414565- 4572
PubMed
Smith  I Mycobacterium tuberculosis pathogenesis and molecular determinants of virulence. Clin Microbiol Rev 2003;16463- 496
PubMed
Kumar  MVNagineni  CNChin  MSHooks  JJDetrick  B Innate immunity in the retina: toll-like receptor (TLR) signaling in human retinal pigment epithelial cells. J Neuroimmunol 2004;1537- 15
PubMed

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