1 Laboratoire de Biologie de la Reproduction, Hôpital Nord, 42055 Saint Etienne, 2 Laboratoire de Génétique Moléculaire, Faculté de Médecine, Saint Etienne and 3 Département de Médecine de la Reproduction, Hôpital Edouard Herriot, Lyon, France
4 To whom correspondence should be addressed. e-mail: rachel.levy{at}chu-st-etienne.fr
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Abstract |
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Key words: France/male infertility/microdeletions/survey/Y chromosome
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Introduction |
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Y chromosome microdeletion screening is of aetiological and prognostic interest. In addition, it can improve genetic counselling.
Improvements in molecular biology techniques have, in theory, made this diagnosis more accessible in routine male infertility analysis. However, not all centres performing IVF have in fact been diagnosing Y chromosome microdeletions.
Our survey sought to establish an overview of French practice with respect to prescribing Yq deletion screening and molecular diagnostic methods, and to answer the following points. (i) How many practitioners and IVF centres request Y chromosome microdeletion screening and what are the prescribing practitioners specialities? (ii) What are the indications adopted? (iii) How is the screening financed? (iv) How many molecular genetic diagnosis laboratories screen for microdeletions? (v) How many analyses are performed? (vi) What methods are used? (vii) How is the analysis invoiced?
Our objectives are to give a (technical, financial) overview of Y chromosome microdeletion screening issues.
Two questionnaires were drawn up, one for prescribers and the other for molecular biologists. These questionnaires were sent out in June 2001 via the persons in charge (clinicians and biologists) in the 88 authorized IVF centres in France (according to the French national FIVNAT register). Survey objectives were explained in an accompanying letter. The persons in charge in these centres (clinicians and biologists) were asked to pass on the questionnaires to prescribers/molecular biologists performing this analysis in or for their centre if they did not have sufficient data to reply themselves or if various prescribers were not in agreement. A second copy of the questionnaire was faxed to those centres that had not replied within 6 months. Physicians who had not replied were contacted by telephone 8 months later. CHUs (teaching hospitals), CHGs (general hospitals), non-profit making establishments and Institutes were classified as public, giving a total of 39 such public centres (44%). Professional establishments, classified as private, were made up of 47 centres (53%). Two entities were classified as mixed as they combined both public (e.g. biology) and private (e.g. gynaecology) activity.
Statistical comparisons were made using the 2-test for qualitative variables.
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Response rate |
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Molecular biologists
From the IVF centre responses, 30 laboratories performing molecular biology analysis were identified. Twenty-nine of these laboratories (97%) answered our questionnaire.
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Reasons for not proposing Y chromosome microdeletion analysis |
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Questions about usefulness
It is worth recalling that the diagnosis of a Y chromosome microdeletion does, as a matter of fact, affect patient management, because freezing of sperm for preventive purposes has been proposed since 2000 (Krausz et al., 2000) on the basis of articles reporting a decreased sperm count in patients with AZFc deletions (Girardi et al., 1997
; Simoni et al., 1997
; Chang et al., 1999
; Saut et al., 2000
; Dada et al., 2003
). Moreover, a positive diagnosis of microdeletion is of prognostic value for the indications for testicular biopsy according to the type of microdeletion found (Brandell et al., 1998
; Silber et al., 1998
; Krausz et al., 2000
; Hopps et al., 2003
), or for possible surgical intervention for varicocele (Cayan et al., 2001
). In close to one-quarter of responses, the non-prescribers pointed out the difficulty of genetic counselling after the discovery of Y chromosome microdeletion; however, the genetic counselling would be better adapted: the phenotype of a son conceived by ICSI cannot yet be definitively predicted, but the deletion is necessarily transmitted (Kent-First et al., 1996
; Mulhall et al., 1997
; Jiang et al., 1999
; Kamischke et al., 1999
; Kleiman et al., 1999
; Cram et al., 2000
; van Golde et al., 2001
; Oates et al., 2002
; Peterlin et al., 2002
). Furthermore, preventive therapy (sperm cryoconservation for successive assisted reproductive techniques) could be proposed to affected sons, as suggested by previous studies (Krausz and McElreavey, 1999
; Toth et al., 2001
; Dada et al., 2003
).
It should also be mentioned that the detection of nullisomic gametes for the sex chromosomes in patients with microdeletions (Siffroi et al., 2000; Jaruzelska et al., 2001
) may lead to the conception of embryos presenting Turners syndrome or sexual ambiguity. The converse has also been demonstrated recently: a Y chromosome microdeletion in AZFc has been reported in 33% of subjects presenting a Turner-like phenotype with sexual ambiguity and a 45,X/46,XY karyotype (Patsalis et al., 2002
). However, there is no single equivalent case among ICSI children of Y-deleted men. Thus, these theories are still controversial.
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Prescribing centres |
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Laboratories performing this analysis |
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Fourteen laboratories carried out a preliminary investigation with a limited number of markers and, in a second step, refined the deletion limits and extent with a series of supplementary markers. The number of markers used for first intention screening, and in second intention, to determine the deletion extent, varied greatly from one centre to another (Figure 5). Eight laboratories performed the diagnosis without specifying the extent of the microdeletion detected.
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Control genes used also greatly varied from one centre to another: the SRY gene was used in 22 centres and the ZFY gene in seven centres. One centre used an autosomal gene (not specified) and another used DAZLA. Two centres made use of an X chromosome gene. The extent to which control subjects were used varied greatly from one laboratory to another (Figure 6): using normal male DNA as reactivity control was not systematic (22/28). Two laboratories did not use a negative control (tube without DNA). Furthermore, the use of reaction specificity controls (womens DNA, or DNA from microdeleted men) varied greatly according to laboratory. Thus, many of the laboratories are not using appropriate negative and positive controls.
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In parallel, the idea of quality control was warmly welcomed. Laboratories would agree to participate in an international (17/29), national (16/29) or both national and international (8/29) quality control scheme. As an example of a quality control programme, the External Quality Assessment, under the auspices of the EAA and the European Molecular Quality Network, represents a real effort being made towards an improvement of Y chromosomal microdeletions diagnosis.
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Results |
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Cost considerations
This analysis is charged to patients at an average rate of 164 euros (range 54635) (Table II). The high cost of the test (43% of responses), with no national health insurance (Sécurité Sociale) cover, is the second most often stated reason for non-prescription. Apart from in research protocols, which are usually of limited duration, the cost of this analysis was borne by the patient (in the private sector) or the hospital (in the public sector). The great variability in cost rating from one centre to another is the result of the overall problem of inadequate financial cover for molecular biology tests under the French system. For this reason, each centre adopts its own policy. Recognition and cover by the Sécurité Sociale appears to be an essential step towards its adoption in French routine practice. The present survey suggests a double cost rating for this analysis: one for detection and one for definition of the extent of Y chromosome microdeletion.
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Conclusions |
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The heterogeneity of the observed protocols calls for standardization, which is an important issue that can be reached only by a joint effort of experts in the field. We call for increased participation of laboratories performing AZF diagnostics in a quality control program, such as the External Quality Assessment.
The current data concerning the French practice should be discussed among experts in this field, who can draw up a consensus concerning screening indications and move towards identifying the most cost effective methods. In France, the current data could also help in the consideration of financial management by health insurance organization (Sécurité Sociale).
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Acknowledgements |
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References |
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Submitted on July 18, 2003; resubmitted on November 25, 2003; accepted on December 3, 2003.
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