© The Author 2006. Published by Oxford University Press.
CORRESPONDENCE |
Re: Evidence for an Association Between Chlamydia psittaci and Ocular Adnexal Lymphomas
Affiliations of authors: Department of Tumor Biology (PD, AV-S, MT, M-CE), Department of Clinical Hematology (DC, DD), Department of Ophthalmology (PA, LL-LR), Department of Radiation Oncology (R. Dolcetti), Institut Curie, Paris, France; Unité de Biologie des Interactions cellulaires, URA CNRS 2582, Institut Pasteur, Paris, France (AS); Department of Oncology (AJMF), Department of Pathology (MP), San Raffaele H Scientific Institute, Milan, Italy; Immunovirology and Biotherapy Unit, Department of Pre-clinical and Epidemiological Research, Centro di Riferimento Oncologico IRCCS National Cancer Institute, Aviano, Italy (FS, R. Dendale); Inserm/Institut Curie U 509, Paris, France (M-HS)
Correspondence to: Didier Decaudin, MD, PhD, Service d'Hématologie Clinique, Institut Curie, 26 rue d'Ulm, 75.248 Paris cedex 05, France (e-mail: didier.decaudin{at}curie.net).
Ferreri et al. (1) recently reported that patients with ocular adnexal lymphoma had a high prevalence of Chlamydia psittaci infection in both tumor tissues and peripheral blood mononuclear cells. More recently, one study reported a similar prevalence of the infection in a Korean patient series (2), whereas three other studies did not observe such an association (3–5).
To investigate the reasons for these different results, our two laboratories, one in Milan (which produced the Ferrari et al. manuscript) and one in Paris, investigated both in independent blinded duplicate the presence of C. psittaci, C. trachomatis, and C. pneumoniae DNA in tumor samples from ophthalmologic biopsies obtained from 16 French patients with ocular adnexal lymphoma and from tissue of two control subjects with other types of lymphoproliferative disease. All 16 patients had histologically proven non-Hodgkin lymphoma of the ocular adnexa (10 at conjunctival sites and six at intraorbital sites). Ten samples from tumor lymph node biopsies from patients diagnosed with marginal zone B-cell lymphoma (n = 8), splenic-type marginal zone B-cell lymphoma (n = 1), and follicular lymphoma (n = 1) were also obtained; 10 nodal biopsy samples from patients diagnosed with reactive lymphoid hyperplasia were also collected.
DNA was extracted from three 15-µm thick sections of paraffin-embedded biopsies that had been fixed in alcohol, formalin, and acetic acid, using the QiampDNA mini kit (Quiagen, Courtaboeuf, France). A multiplex touchdown enzyme time-release polymerase chain reaction (PCR) was performed that was designed to simultaneously detect the DNA sequences of C. psittaci, C. pneumoniae, and C. trachomatis. Briefly, 10 mL of template DNA were used in 40 mL of PCR mix that included 25 pmol of each primer (1,6), 0.25 mM deoxynucleotide triphosphates, and 2 U of AmpliTaq Gold polymerase (Applied Biosystems, Foster City, CA) in a 2.5 mM MgCl2 buffer. Blank reactions were inserted between samples to rule out contamination between samples. A mix of DNA extracted from cells infected by each of the three chlamydial species was included as a positive control. To determine the limit of detection of the PCR, we cloned each PCR amplicon into a plasmid that we used as a template. We were able to detect as few as 10 copies of template per sample. A second set of PCRs, using a human reference gene (GAPDH) (6), was used to ensure the quality of the PCR and to prove that no PCR inhibitors were present in any sample. PCR analyses were performed in duplicate in our two independent laboratories.
C. psittaci DNA was detected in the tumor tissue of only one patient with follicular ocular adnexal lymphoma. No C. psittaci, C. pneumoniae, or C. trachomatis DNA sequences were detected in any of the tumor samples obtained from other patients diagnosed with ocular adnexal lymphoma, nodal lymphoma, or lymphoid hyperplasia.
In conclusion, the prevalence of C. psittaci infection in this series of ocular adnexal lymphoma patients was considerably lower than that reported by Ferreri et al. (1). The identical results obtained by our two laboratories indicate that this finding is not due to different experimental conditions. It could be explained by a heterogeneous distribution of the bacterial infection. However, objective responses observed after C. psittaci eradication with doxycycline suggest the role of this microorganism in the maintenance of ocular adnexal lymphoma (7).
REFERENCES
(1) Ferreri AJ, Guidoboni M, Ponzoni M, De Conciliis C, Dell'Oro S, Fleischhauer K, et al. Evidence for an association between Chlamydia psittaci and ocular adnexal lymphomas. J Natl Cancer Inst 2004;96:586–94.
(2) You C, Ryu M, Huh J, Park J, Kang H, Ahn H. Ocular adnexal lymphoma is highly associated with Chlamydia psittaci. Eur J Cancer 2005:282–3.
(3) Rosado MF, Byrne GE Jr, Ding F, Fields KA, Ruiz P, Dubovy SR, et al. Ocular adnexal lymphoma: a clinicopathological study of a large cohort of patients with no evidence for an association with Chlamydia psittaci. Blood 2005;107:467–72.
(4) Gracia E, Mazzucchelli L, Frösch P, Jimenez J, Rodriguez D, Capo V, et al. Low prevalence of Chlamydia psittaci infection in ocular adnexal lymphoma from Cuban patients [abstract 303]. Ann Oncol 2005;16.
(5) Vargas RL, Fallone E, Felgar RE, Feiedberg JW, Arbini AA, Andersen AA, et al. Is there an association between ocular adnexal lymphoma and infection with Chlamydia psittaci? The University of Rochester experience (October 21, 2005). Leuk Res (DOI: 10.1016/j.leuk.res.2005.09.012).
(6) de Cremoux P, Thioux M, Peter M, Vielh P, Michon J, Delattre O, et al. Polymerase chain reaction compared with dot blotting for the determination of N-myc gene amplification in neuroblastoma. Int J Cancer 1997;72:518–21.[CrossRef][ISI][Medline]
(7) Ferreri AJ, Ponzoni M, Guidoboni M, De Conciliis C, Resti AG, et al. Regression of ocular adnexal lymphoma after Chlamydia psittaci-eradicating antibiotic therapy. J Clin Oncol 2005;23:5067–73.
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