REPRODUCTIVE ENDOCRINOLOGY
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1 REPRODUCTIVE ENDOCRINOLOGY FERTILITY AND STERILITY VOL. 72, NO. 5, NOVEMBER 1999 Copyright 1999 American Society for Reproductive Medicine Published by Elsevier Science Inc. Printed on acid-free paper in U.S.A. Efficacy of methods for determining ovulation in a natural family planning program Maurizio Guida, M.D., Giovanni A. Tommaselli, M.D., Stefano Palomba, M.D., Massimiliano Pellicano, M.D., Gianfranco Moccia, M.D., Costantino Di Carlo, M.D., and Carmine Nappi, M.D. Department of Obstetrics, Gynecology and Physiopathology of Reproduction, University of Naples Federico II, Naples, Italy Objective: To evaluate the efficacy in ovulation detection of methods used in natural family planning in comparison with pelvic ultrasonography. Design: Prospective analysis of ovulation detection by natural family planning methods and ultrasonography. Setting: Natural family planning clinic, Department of Obstetrics and Gynecology, University of Naples Federico II. Patient(s): Forty healthy women who were highly motivated to use natural family planning. Intervention(s): None. Main Outcome Measure(s): Transvaginal ultrasonographic findings, urinary LH levels, salivary -glucuronidase activity, salivary ferning levels and characteristics of cervical mucus, and BBT. Result(s): Urinary LH level determination yielded a 100% correlation with the simultaneous ultrasonographic diagnosis of ovulation. Mucus sensations and characteristics yielded a 48.3% correlation when simultaneously evaluated with ovulation. -Glucuronidase levels yielded a 27.7% correlation. The salivary ferning test had a 36.8% ovulation-detection rate the day of ovulation, but 58.7% of results were uninterpretable. Body temperature measurements yielded a 30.4% correlation with the simultaneous ultrasonographic diagnosis of ovulation. Conclusion(s): Measuring urinary LH levels is an excellent method for determining ovulation. Although variations in mucus characteristics and basal body temperature correlate somewhat with ovulation, the length of the fertile period is overestimated with these methods. The salivary ferning test and measurement of -glucuronidase levels are not good methods for home ovulation testing. (Fertil Steril 1999;72: by American Society for Reproductive Medicine.) Key Words: NFP methods, ovulation self-detection, fertility control Received November 23, 1998; revised and accepted June 18, Reprint requests: Maurizio Guida, M.D., Via U. Ricci 3, Naples, Italy (FAX: ; pellican@unina.it) /99/$20.00 PII S (99) Controlling reproduction is one of the major issues in social economics and preventive medicine today. Limitation of the number of births is essential in underdeveloped countries. Unfortunately, economic problems there will hinder the use of new, expensive hormonal or surgical contraceptive methods in the next decade (1). In developed countries, pharmacologic and instrumental interventions are becoming less accepted, there is a need for nonmedical contraception, and researchers seek to gain a better understanding of physiology; in these countries, the need for nonhormonal, noninvasive methods is well documented (1). New developments in the area of natural family planning have emerged in recent years (1). The World Health Organization (2 6) has played a leading role in the organization of studies for the evaluation of methods for controlling human fertility. At the same time, basic and clinical research are being carried out to discover new technologies that can support, from a scientific point of view, natural methods in family planning. We organized a natural family planning program to support a group of selected couples with information about and training in two sympothermal methods and to evaluate the effectiveness of these two methods. Evaluation 900
2 of the results of this program is still in progress. In addition, we also evaluated single methods of self-detection of ovulation compared with an objective ovulation-determination technique (pelvic ultrasonography). We report the efficacy of these methods in detecting ovulation in the study population. MATERIALS AND METHODS Subjects Forty women were enrolled in the natural family planning program (mean [ SD] age, years; range, years). Exclusion criteria were infertility or subfertility (each women had had a previous pregnancy and had no history of recurrent miscarriage), any medical illness (e.g., diabetes, cardiovascular disease, or gastrointestinal disease), oral contraceptive use in the last 3 months, and any behavior that indicated that the couple was unreliable in terms of following natural family planning rules. To increase the possibility of compliance, we recruited our subjects from groups with high motivation in this area, such as religious groups and university students. Couples were allowed 6 months to become familiar with all of the techniques used in the study. We did not request institutional review board approval, because all patients referred to our clinic had requested natural family planning methods for contraceptive purposes and did not receive any drugs during the study. All women gave informed consent. Ovulation-Detection Methods During each cycle, the following ovulation-detection methods were used: 1. Transvaginal ultrasonography. This technique was performed to determine ovulation objectively. An ultrasonic scanner (SSA250-A; Toshiba, Medical Systems, Rome, Italy) with a 7.5-MHz vaginal probe was used, and the scans were performed daily, starting 7 days after the onset of menses, by the same operator (G.A.T.), who was blinded to the results of the other tests. The day of ovulation was retrospectively identified as the day before a corpus luteum was observed. 2. Daily morning urinary LH level determinations. Levels were determined by the subjects themselves, beginning on day 6 of the cycle. Subjects used a stick system (Clearplan; Farmades, Rome, Italy) based on monoclonal antibodies directed versus LH bound to a colorimetric substance. 3. Determination of salivary -glucuronidase activity. This evaluation was done in our laboratory on specimens that were collected by the subjects starting on day 6 of the cycle and then frozen. The phenolphtalein test and titolation with a colorimetric assay (7) were used. The occurrence of ovulation was hypothesized when the salivary concentration of the enzyme reached, for the first time from the beginning of the cycle, 500 pmol of free phenolphtalein per microgram. 4. The salivary ferning test. Salivary ferning was evaluated daily throughout the cycle with an optical microscope. Dried, unstained specimens were used. A score was assigned; 0 was assigned when salivary ramifications were absent, 1 when there was first-order ferning, 2 when there was second-order ferning, and 3 when there was third-order ferning (8). A score of 1 or 2 was considered indicative of the periovulatory period, and a score of 3 suggested ovulation. 5. Determination of cervical mucus levels and characterization of cervical mucus. The vulva was classified as dry, moist, or wet, and mucus was classified as absent, sticky, or stretchy (9). Identification of the last day in which fluid mucus was observed (the so-called mucus peak (9) was performed retrospectively, considering the last day in which there were a wet, slippery sensation and/or transparent, stretchy mucus. 6. Measurement of rectal or oral BBT. Measurements were done with a digital thermometer (Terumo, Tokyo, Japan) after at least 2 hours of bed rest. Each subject measured BBT at the same site throughout the study. Sexual Activity Couples were instructed to abstain from sexual intercourse as soon as one of the methods indicated the beginning of the fertile period. They were allowed to resume sexual activity only after all methods had indicated the end of the fertile period. To avoid the necessity for prolonged sexual abstinence, the use of a barrier method (i.e., condoms) was permitted. Data Recording Women recorded urinary LH levels, salivary ferning scores, characteristics and sensations at the vulva of cervical mucus, and daily BBTs along with occurrences of discharge of vaginal blood and frequencies of sexual intercourse. All values were entered with ultrasonographic data and -glucuronidase levels onto a spreadsheet program (Excel; Microsoft, Redmond, WA.), to compare the coincidence of the ovulation day determined by each method with the ovulation day determined by ultrasonography. The day of ovulation determined by ultrasonography was considered day 0. If a method indicated ovulation on the same day, simultaneous correlation with ovulation was considered to exist. For each subject, we calculated the percentage of simultaneous correlation for each method in each cycle throughout the study. We then calculated the overall simultaneous correlation for all methods. We also evaluated the presence of indication of ovulation for each method over a length of time, day by day, from 6 days before to 6 days after ovulation as diagnosed by ultrasonography. Statistics To identify significant differences between the methods used for detection of ovulation, we used the instrumental estimation test. With this test, the accuracy and the precision of each method were compared with those of the other methods. The desired outcome was that the method have relatively the same accuracy (as measured by the mean) and the same precision (as measured by the variance). In this study, to compare transvaginal ultrasonography with other methods for detection of ovulation, each method FERTILITY & STERILITY 901
3 FIGURE 1 Timing of ovulation as determined by the different methods throughout a 12-day periovulatory period. } urinary LH kit; -glucuronidase level determination; Œ salivary ferning test; determination of mucus characteristics; BBT measurement. Guida. Determining ovulation. Fertil Steril was used in every cycle. Successively, on a cycle-by-cycle basis, the values from the two methods were added and then subtracted. A statistical test was previously developed by Maloney and Rastogi (10), who showed that sums and differences such as these could be used to detect differences in the variance (precision) of two methods. The F test for comparing variances from two independent samples was not used, because values were obtained in the same cycle from each method; thus, the sample was not independent. Differences in accuracies (mean) were examined with the use of the matched pairs test. RESULTS A total of 148 menstrual cycles were completed by the subjects studied. The mean ( SD) number of cycles per woman was and the median number of cycles per women was 4, with a lower quartile of 3, an upper quartile of 4, and a quartile range of 1. Ultrasonography was performed in all 148 cycles. Urinary LH level determinations were performed in 98 cycles (66%), -glucuronidase level determinations in 95 (64%), salivary ferning observation in 125 (84%), and mucus evaluation and BBT measurements in all 148 cycles (100%). Figure 1 shows the timing of ovulation throughout the cycle as determined by each method. Urinary LH and -glucuronidase levels yielded a 100% and 27.7% correlation, respectively, with the ultrasonographic diagnosis of ovulation. The salivary ferning test had a 36.8% simultaneous correlation with ovulation but had a high percentage (58.7%) of uninterpretable pattern. Mucus sensation and characteristics and body temperature measurements yielded a 48.3% and 30.4% correlating, respectively, when simultaneously evaluated with ovulation. Ovulation timing determined by urinary LH level measurement always coincided with ultrasonographically detected (actual) ovulation measurement of salivary -glucuronidase levels and indicated ovulation 3 days after actual ovulation in 27.7% of cases and 2 days after actual ovulation in 16.6% of cases. The salivary ferning test indicated ovulation the day before actual ovulation in 21% of cases and the day after in another 21%. Determination of mucus sensation and characteristics indicated ovulation the day after actual ovulation in 27.4% of cases. Finally, body temperature measurements indicated ovulation the day before ultrasonographically detected ovulation in 32% of cases. There was no difference in the accuracy and precision of urinary LH level determination and those of transvaginal ultrasonography (P.05). For the other methods, analysis of variance showed that precision differed significantly from that of ultrasonography. Analysis of accuracy (mean) showed that the accuracy of determination of mucus sensation and characteristics was not significantly different from that of transvaginal ultrasonography (t 1.23; P.05, two-tailed). Measurement of salivary -glucuronidase levels (t 3.38) and measurement of basal temperature (t 4.31) are not good methods for determination of ovulation, because their accuracies differ significantly from that of transvaginal ultrasonography (P.05). The salivary ferning test did not significantly differ from ultrasonography in terms of accuracy (t 0.66; P.506); however, 58.7% of results were uninterpretable and thus excluded from analysis. During this study there were two unwanted pregnancies, yielding a Pearl index ([number of pregnancies number of cycles observed]/1,300) (1) of DISCUSSION Natural family planning methods are widely used today. In Peru and Mauritius, nearly one in five women reported using natural family planning methods (11). In 1998, 4% of fertile married women in the United States relied on these methods (12). In Italy, 16% of fertile couples use periodic abstinence as a means of contraception (13). Nevertheless, most of these couples use improper methods to determine the fertile period of the cycle, mostly because of inadequate information or training. It is therefore important to find out which of the methods used in natural family planning is the most effective for determining ovulation. From the data gathered in this study, it is clear that the 902 Guida et al. Ovulation and natural family planning Vol. 72, No. 5, November 1999
4 most accurate method is self-determination of urinary LH levels using a home kit. Another method for determining the fertile period, which seems to be correlated with the periovulatory period, is observation of mucus characteristics. Measurement of salivary -glucuronidase levels cannot be considered reliable for detecting the ovulation day, given that it indicated ovulation both 2 and 3 days after ultrasonographically detected ovulation in significant percentages of subjects. Basal body temperature is capable of indicating the fertile period, even though detected ovulation days may be scattered throughout the periovulatory period, thus inducing extended periods of sexual abstinence. The salivary ferning test proved capable of indicating ovulation, but only after the exclusion of more than half of the results (58.7%), which were uninterpretable. The effectiveness of measurement of urinary LH levels for the purpose of monitoring ovarian activity and detecting ovulation is well documented and was first confirmed in the area of infertility. Martinez et al. (14) evaluated the results of IUI timed either after the detection of a urinary LH surge at home by the patients or after a positive LH test interpreted by a gynecologist. The authors concluded that a home urinary LH test was a reliable device for the prediction of ovulation, because the patients and gynecologist s results agreed in 89% of cases. A similar study was conducted by Robinson et al. (15), who tried to determine whether a home urinary LH detection method could predict ovulation and thus decrease the number of controls required for the management of a donor insemination cycle. They observed that the use of such a method reduced the number of visits required and did not worsen monthly fecundity or cumulative conception rate. The ability of home urinary LH level determination to detect ovulation in spontaneous and induced cycles has also been tested (16). In that study, the relationship between urinary LH levels, serum LH levels, serum E 2 levels, and ultrasonography was examined. The results indicated that urine LH level determination was a good method of predicting ovulation in spontaneous cycles. In a previous study, our group evaluated the results of the use of two commercial immunoenzymatic kits for the determination of urinary LH levels comparing results with ultrasonographic findings regarding ovulation (17). Kit A was a qualitative assay and kit B was quantitative and monophasic. The first had a sensitivity of 30%, a specificity of 99%, and an accuracy of 92%, whereas the second had a sensitivity of 80%, a specificity of 90%, and an accuracy of 88%. It is evident that these results are in agreement with the literature. The only difference found was a slightly higher accuracy, which can be explained both by higher motivation in the group and by the fact that not all of the subjects used kits in all the cycles, which may have affected the overall simultaneous correlation. Among the other methods evaluated in the current study, only determination of mucus characteristics showed good correlation with ultrasonography in terms of diagnosis of ovulation. Billing s method (or the ovulation method), based on the observation of the modification of cervical mucus throughout the cycle, is one of the main natural family planning methods used and its efficacy has been proved in several studies [see Guida et al. (1) for a review]. There is a high degree of skill involved both in teaching and in using the method. Because patients must be skilled and dedicated to use this method, it is highly effective. Determination of BBT in the present study did show a slight correlation with ultrasonography with regard to diagnosis of ovulation. This finding is in contrast with findings by Martinez et al. (14). These investigators found a truepositive rate of 90% and a false-negative rate of 2%; 8% of readings were uninterpretable, reflecting measurement problems. They also found that the thermal nadir occurred within 1 day of urinary LH surge in 75% of cases and in 90% when 2 days were considered. This discrepancy can be explained by the fact that BBT measurement has been proved effective for detecting fertile periods if performed in association with observation of other signs and symptoms of ovulation and has been shown to indicate only the periovulatory period. Indeed, in 94% of cases, BBT-determined ovulation days were scattered from day 1 to day 3 of actual ovulation, giving a fairly good indication of the fertile period but inducing prolonged sexual abstinence. The salivary ferning test had a good percentage of simultaneous correlation, but the high percentage of uninterpretable results must be considered as well. Most of the records did not show the characteristic triphasic pattern reported in the literature (8), but they frequently showed a monophasic pattern. This may be due to the inconstant estrogen dilution in the saliva, yielding different patterns between women and within the same subject between cycles. This discrepancy may also be due to the fact that the slides were interpreted by the subjects themselves only, not also by a gynecologist, who would be more skilled in interpreting slides; as a result, there was a higher percentage of uninterpretable results. It is evident from these data that salivary ferning observation by natural family planning users cannot yet be relied on for accurate identification of the fertile period. There are very few published data regarding the method of salivary -glucuronidase level determination. The pattern of this enzyme throughout the cycle has been found to be triphasic, with the second peak coincident with ovulation (7). In the present study, however, we observed a biphasic pattern, with a lower peak of coincidence on ovulation day and a higher peak on day 3. Women may not recognize the first peak and may thus have unprotected sexual intercourse in the fertile period. These data seem to indicate that determination of levels of this salivary enzyme cannot be regarded as a method for self-identification of ovulation and these findings confirm the unreliability of salivary enzyme levels as indicators of ovarian activity. FERTILITY & STERILITY 903
5 We believe that the low Pearl index in this study is not indicative, because our study involved a limited number of motivated women and a restricted number of cycles. We are currently conducting a study involving a larger number of cycles and are evaluating these methods combined into a modified sympothermal method. The cost of urinary LH kits is an important issue, given that one of the aims of natural family planning is to decrease medical expenses. The prices of LH kits are still high (approximately $40 $45 per cycle in Italy), but use of natural methods as well to determine ovulation should help reduce the number of determinations per cycle, thus limiting costs. Moreover, women with regular cycles can use alternative methods (determination of mucus characteristics, BBT measurement) to determine the period of increased fertility, limiting the need for LH level determinations to a few days per cycle. Furthermore, the use and number of LH level determinations are also linked to the acceptable period of sexual abstinence. Couples who wish to have the shortest period of sexual abstinence will need to monitor urinary LH levels accurately. On the other hand, subjects who accept a longer period of sexual abstinence can rely also on other methods that indicate the beginning and end of the fertile period. In conclusion, this study seems to confirm that selfdetermination of urinary LH levels is a reliable way to determine ovulation. This method, in association with other fertility indices, minimizes the period of sexual abstinence necessary for couples using natural family planning methods and indicates the right days to have sexual intercourse for those desiring pregnancy. Furthermore, although variations in mucus characteristics and BBT have a certain degree of coincidence with ovulation, the length of the fertile period is overestimated with these methods. Finally, the salivary ferning test and measurement of -glucuronidase levels are not accurate methods for self-determination of ovulation. Acknowledgments. The authors thank Sergio Scippacerola, Ph.D., Department of Mathematics and Statistics, University of Naples Federico II, for his substantial collaboration in the statistical analysis. References 1. Guida M, Tommaselli GA, Pellicano M, Palomba S, Nappi C. An overview of the effectiveness of natural family planning. Gynecol Endocrinol 1997;11: A prospective multicentre trial of the ovulation method of natural family planning. I. The teaching phase. Fertil Steril 1981;36: A prospective multicentre trial of the ovulation method of natural family planning. II. The effectiveness phase. Fertil Steril 1981;36: A prospective multicentre trial of the ovulation method of natural family planning. III. Characteristics of the menstrual cycle and of the fertile phase. Fertil Steril 1983;40: A prospective multicentre trial of the ovulation method of natural family planning. IV. The outcome of pregnancy. Fertil Steril 1984;41: A prospective multicentre trial of the ovulation method of natural family planning. V. Psychosexual aspects. Fertil Steril 1987;47: Bardin GW, Brown TR, Mills NC, Gupta C, Bullock LD. The regulation of beta-glucuronidase gene by androgens and progestins. Biol Reprod 1978;18: Guida M, Barbato M, Bruno P, Lauro G, Lampariello C. Salivary ferning and the menstrual cycle in women. Clin Exp Obstet Gynecol 1993;20: Billings JJ (ed.). The ovulation method. Melbourne: Advocate Press, Maloney CJ, Rastogi SC. Significance tests for Grubbs estimators. Biometrics 1970;26: Stanford JB, Lemaire JC, Thurman PB. Women s interest in natural family planning. J Fam Pract 1998;46: Forrest JD, Fordyce RR. U.S. women s contraceptive attitudes and practice: how have they changed in the 1980s? Fam Plann Perspect 1988;20: Italian National Council for Research. Fertility survey in Italy. General report. Vol. 1. Italian National Council for Research, Martinez AR, Bernardus RF, Vermeiden JP, Schoemaker J. Reliability of home urinary LH tests for timing of insemination: a consumer s study. Hum Reprod 1992;7: Robinson JN, Lockwood GM, Dalton JD, Franklin PA, Farr MM, Barlow DH. A randomized prospective study to assess the effect of the use of home urinary luteinizing hormone detection on the efficiency of donor insemination. Hum Reprod 1992;7: Kawano T, Matsuura K, Honda R, Nishimura H, Tanaka N, Okamura H. Prediction of ovulation by urinary LH surge [in Japanese]. Nippon Naibunpi Gakkai Zasshi 1992;68: Cardone A, Guida M, Lampariello C, Bruno P, Montemagno U. Objective and subjective data for fertile period diagnosis in women: comparison of methods. Clin Exp Obstet Gynecol 1991;19: Guida et al. Ovulation and natural family planning Vol. 72, No. 5, November 1999
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