Institute of Reproductive Medicine of the University, Domagkstrasse 11, D-48129 Münster, Germany
Deception by chimerism
Dear Sir,
With great interest we read the debates and articles on the molecular diagnosis of Y chromosomal microdeletions (Pryor and Roberts, 1998; Simoni et al., 1998
; Vogt, 1998
). We recently had an interesting case adding an unexpected aspect to the question of quality control.
Infertility affects ~7% of the male population. There is increasing evidence for a genetic cause for at least a subset of fertility disorders and DNA analysis is being introduced more and more into the routine infertility work-up. A set of spermatogenesis-specific genes are known to be involved in fully normal germ cell proliferation and recently microdeletions of the Y chromosome have been detected in 510% of men with idiopathic azoospermia and oligozoospermia. According to Vogt et al. (1996), the microdeletions can be subgrouped into the AZF (azoospermia factor) a to c region. Candidate genes, e.g. RBM or DAZ, can be allocated to these regions.
A 53 year old patient presented with the questions of his chances of fertility prior to his second marriage. He had an adult daughter from a previous relationship. Semen analysis revealed azoospermia. He had been treated for chronic myeloic leukaemia (Philadelphia chromosome positive) 3 years ago. Total body irradiation and chemotherapy associated with autologous bone marrow transplantation in most cases lead to complete and irreversible spermatogenic failure, through sporadic recovery of the reproductive axis has been reported.
Factors that determine the fate of spermatogenesis after oncological therapy are not well understood (Chatterje et al., 1996). To further elucidate the pathogenesis of azoospermia and provide better prognostic information to the patient, we performed Y chromosomal microdeletion analysis, which is normally included into the diagnostic work up of men presenting with severe impairment of spermatogenesis in our infertility clinic. We routinely use nine different Y chromosomal markers and one autosomal marker to ensure proper polymerase chain reaction (PCR) conditions.
In our patient, amplification of Y chromosomal markers for the different AZF regions failed completely, however, the autosomal marker could be amplified by PCR from genomic DNA obtained from leukocytes. In subsequent steps we tried to amplify SRY, a crucial marker for the testis formation, which was also absent in this patient. The lack of any specific male marker in the genomic DNA from the blood led us to speculate on the sex of the bone marrow donor, which indeed turned out to be female. Analysis of DNA isolated from buccal cells of the patient then showed the normal presence of all Y chromosomal markers.
Molecular diagnostics on genomic DNA isolated from lymphocytes is becoming an important tool in many different clinical conditions. In infertile males, this diagnostic procedure is now being performed world-wide using sequenced-tagged site (STS) markers covering the AZF areas (Simoni et al., 1997). Physicians and laboratories should be aware of the fact that chimerism is created by transplantation centres, which in our case lead to a laboratory deception in the analysis on Y chromosome deletions. We suggest that SRY, as a male marker, should be included into DNA diagnostic of infertile men to assure sex identity.
References
Chatterje, R. and Goldstone, A.H. (1996) Gonadal damage and effects on fertility in adult patients with hematological malignancy undergoing stem cell transplantation. Bone Marrow Transpl., 17, 511.[ISI][Medline]
Pryor, J.L. and Roberts, K.P (1998) Principles of sequence-tagged site selection in screening for Y deletions. Hum. Reprod., 13, 1768.[ISI][Medline]
Simoni, M., Gromoll, J., Dworniczak, B. et al. (1997) Screening for deletion of the Y chromosome involving the DAZ (Deleted in Azoospermia) gene in azoospermia and severe oligozoospermia. Fertil. Steril., 67, 542547.[ISI][Medline]
Simoni, M., Kamischke, A. and Nieschlag, E. (1998) Initiative for international quality control. Hum. Reprod., 13, 17641768.
Vogt, P.H. (1998) Human chromosome deletions in Yq11, AZF candidate genes and male infertility: history and update. Mol. Hum. Reprod., 4, 739744.[Abstract]
Vogt, P.H., Edelmann, A., Kirsch, S. et al. (1996) Human Y chromosome azoospermia factors (AZF) mapped to different subregions in Yq11. Hum. Mol. Genet., 5, 933943.