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Seasonal variation in spermatogenesis in the nine-banded armadillo (Dasypus novemcinctus) from Southeastern Brazil
Animal Reproduction Science, 6 (1983) 135--141
135
Elsevier Science Publishers B.V., Amsterdam -- Printed in The Netherlands
SEASONAL VARIATION IN SPERMATOGENESIS IN THE NINE-BANDED ARMADILLO (DASYPUSNOVEMCINCTUS)FROM SOUTHEASTERN BRAZIL
C.N. T O R R E S , H.P. G O D I N H O
and A.B.M. M A C H A D O
Department of Morphology, Institute of Biological Sciences, Federal University of Minas Gerais, Belo Horizonte, M G (Brazil) (Accepted 27 M a y 1983)
ABSTRACT
Tortes, C.N., Godinho, H.P. and Machado, A.B.M., 1983. Seasonal variation in spermatogenesis in the nine-banded armadilI~ (Dasypus novemeinctus) from southeastern Brazil. Anita. Reprod. Sci., 6: 135--141. Nine-banded armadillos living in the southeastern region of Brazil are seasonal breeders. The weight of the testes, diameter of seminiferous tubules, the number of type A spermatogonia and round spermatids at stage 1 of the cycle of the seminiferous epithelium, and volumetric proportions of Leydig cells showed significant variations during the annual reproductive cycle. These parameters reached a peak during the months of November and December when rainfall was plentiful and the number of daily hours of sunlight was a t its highest level. In the period of June to August the above parameters were at their lowest values.
INTRODUCTION
In a preliminary paper, Godinho et al. (1977) demonstrated a yearly variation in testis and seminal vesicle weights and in citric acid and fructose contents of the seminal vesicles of nine-banded armadillos. Czekala et al. (1980) have shown in nine-banded armadillos from the Northern hemisphere that the highest concentration of plasma testosterone occurred during the summer and that it was maintained at relatively high levels during the other seasons, indicating that testicular function continued throughout the year. They also reported that the testicularhistology of these animals was identical at all times of the year, in agreement with previous works by Weaker (1977a, 1977b), who detected no major seasonal modifications in the spermatogenic process nor in the interstitialtesticular tissue of the nine-banded armadillo. Recently, Peppler and Stone (1981) have also demonstrated that the plasma testosterone level of the same species does not significantly change during the year although it is higher in the summer than in the winter. In the present paper, histometrical analyses of testicular structures from
0378-4320/83/$03.00
© 1983 Elsevier Science Publishers B.V.
136
nine-banded armadillos living in southeastern Brazil were performed in order to evaluate the morphological changes that may occur in the testes during the annual reproductive cycle. MATERIALS AND METHODS
Thirty-one male nine-banded armadillos, 3.5--4.2 kg in body weight, were captured during the period of one year (2 animals were captured in each of the months Sep., Oct., Apr., July and Aug., and 3 in each of Nov., Dec., Jan., Feb., Mar., May and June), near Belo Horizonte (19°49 ~S). After killing the animals, their testes were removed from the abdominal cavity and then weighed (data for testes weight indicate the mean weight of right and left testes). Fragments of the testes were fixed in Susa's fluid, embedded in paraffin wax and sectioned at 10 pm. Hematoxylin-eosin staining was used throughout the study. The nuclei of type A spermatogonia, round spermatids and Sertoli cells were counted in 10 randomly selected cross-sections of seminiferous tubules at stage 1 of the cycle of the seminiferous epithelium in each animal. The diameters of these cross-sections were measured. Stage 1 encompasses the portion of the cycle extending from the shedding of the spermatozoa into the lumen of the seminiferous tubule to the beginning of spermatid nuclei elongation. This corresponds to the first stage of the cycle in the eight-stage method of Roosen-Runge and Giesel (1950) for the rat and applied to the nine-banded armadillo (Tortes et al., 1981). The single generation of spermarids present in stage 1 constitutes the group of most advanced germinal cells in the cycle; this allows determinations of their number with ease. Volumetric proportions, expressed in percentages, of testis structures were determined according to Elias et al. (1971) with the use of a 25-point ocular integrator. The coincidences between the ocular grid points and Leydig ceils, seminiferous tubule wall, tubular lumen and the other intertubular structures were recorded as obtained from 200 counts per animal. The least significant mean difference test (Snedecor and Cochran, 1967) was used to compare the monthly variations among testicular structures (P < 0.05). RESULTS
Testis weight. Testes attained their maximal weight of 8.8 +- 1.7 g in November; they decreased in weight from December until June when they reached 4.4 -+ 1.8 g (Fig. 1,F). The testes of animals killed in September and November were significantly heavier than those killed in May, June, July and August. Diameter of seminiferous tubules. The diameter of the seminiferous tubules reached the maximal value of 230 + 18.9 pm in November and decreased
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Fig. 1. Seasonal changes in testes weight (solid circles) and seminiferous tubule diameter (open circles) (F), number of spermatogonia (solid circles) and round spermatids (open circles) in stage 1 of the cycle of the seminiferous epithelium (E) and volumetric proportion, expremed in percentage, of Leydig cells (D) of nine-banded armadillos as related to daily hours of sunlight (C), temperature (B) and rainfall (A) during the period of one year near Belo Horizonte, Brazil (19°49'S); gradually to 161 + 20.3 ~m in June (Fig. 1,F). The diameter of the seminiferous tubules in November was significantly greater than those o f May, June, July and August and in December it was significantly greater than in June and August (Figs. 2,3).
Type A spermatogonia. The highest number (7.5 -+ 3.5) o f type A spermatogonia per cross~ection o f seminiferous tubules at stage 1 o f the cycle was observed in November; in the other months their number was about half of that o f November (Fig. 1,E). The number of type A spermatogonia in November was significantly greater than that of any other month.
138
Fig. 2, 3. Transverse sections of seminiferous tubules of armadillos at stage 1 of the cycle of the seminiferous epithelium (asterisks) taken at the periods of maximal (December, Fig. 2) and minimal (June, Fig. 3) spermatogenic activity in the annual reproductive cycle. H--E, 192X.
Round spermatids. The number of round spermatids (Fig. 1,E) in stage 1 of the cycle reached a peak of 259 + 65 in December. Their number gradually decreased to reach 141 + 21 in August. The number of such spermatids f o u n d in November and December was significantly greater than those of the periods o f May to June and May to August, respectively (Figs. 2,3).
139
Sertoli cells. The number of Sertoli cells did not show appreciable variation during the cycle.
Volumetric proportions of testis structures. Leydig cells were the intertubulax structures which showed the widest range of variation during the annual cycle (Fig. 1,D). They reached a peak of 11 + 1.3% in November and gradually decreased until June when they reached 2.2 + 0.7% of the testis volume. The November value was significantly greater than those of the months of February to August. The other components of the intertubular space as well as tubular lumen and tubular wall did not present marked volumetric variations during the year. Climatic parameters. Rainfall, maximal and minimal mean air temperature and hours of sunlight occurring in the region during the period of study are presented in Fig. 1 (A, B, C). DISCUSSION
An analysis on some aspects of the reproductive biology of nine-banded armadillos (Dasypus novemcinctus) of the Southern hemisphere as compared to those of the Northern hemisphere is now presented. The various histometrical parameters of the testis of the nine-banded armadillos studied in the present work showed significant annual variation (except for the Sertoli cells), in disagreement with the histological findings of Weaker (1977a, 1977b) and Czekala et al. (1980). The values for those parameters reached a peak during the months of November and December (summer in the Southern hemisphere). At this time of the year, rainfall was plentiful and the number of daily hours of sunlight was at its highest. At a corresponding season, nine-banded armadillos from the Northern hemisphere show peak testicular activity (Czekala et al., 1980; Peppler and Stone, 1981). During the period of June to August those testis parameters of the Brazilian nine-banded armadillos were at their lowest values in the annual cycle. This was the time when rainfall was almost zero and the daily hours of sunlight markedly diminished. The annual reproductive cycle of females of ninebanded armadillos has been studied mainly in specimens living in the Northern hemisphere (Newman, 1913; Hamlett, 1932; Enders, 1966; Peppler and Canale, 1980). In these animals, ovulation and fertilization occur from June to August; retarded implantation of the blastocyst takes place in November or December and is followed by a gestation period of about 4 months; birth of monozygous quadruplets occurs in February or March. The only information available on the reproductive cycle of female nine-banded armadillos in Brazil consists of unpublished data of one (ABMM) of the authors of the present paper, who found that females captured in December, in the same area as those of the present study, showed ovaries with mature follicles or corpora lutea, and the fundus of the uterus more vascularized than the rest
140
of the organ. In June, the uterus contained fetuses 1.6--2.0 cm in length; in August the fetuses measured about 8.5 cm. The young were born in September. As observed in several other wild mammals (Sadleir, 1969), a change in the reproductive cycle, associated with the inversion of the seasons of the year, is apparent in nine-banded armadillos living in the Southern hemisphere as compared with those from the Northern hemisphere. Photoperiod, temperature and rainfall are usually related to the beginning of the breeding season. Near the Equator, the very small variation in the photoperiod seems to have no effect on mammalian reproduction (Sadleir, 1969). At the latitude of the area under study, 20°S, the increase of about 1 rain day-1 in the photoperiod, between the June and December solstices, may be sufficient to affect the reproductive cycle in nine-banded armadillos. It is difficult to perceive any relationship between temperature and reproduction in our nine-banded armadillos. The maximal daily mean temperature in our region does not change considerably, remaining at around 28-32°C throughout the year, whereas the minimal mean temperature falls to around 4°C during the dry season. Over this temperature range it is possible that nine-banded armadillos may be affected only in their search for food, limiting it to the more comfortable hours of the day, as indicated by Taber (1945). There seems to be a positive correlation between minfaU and the breeding season of the nine-banded armadillo. Immediately after the first rains the availability of food increases and, as observed in mammals of Panama (Flemming, 1973), this time coincides with the weaning period. Mating would occur shortly thereafter. The available data on .the reproductive cycle of nine-banded armadillos living near Belo Horizonte suggest that these animals are seasonal breeders. Spermatogenic activity is maximal in November and December, within the rainy season, and then it progressively decreases until June, without, however, reaching a complete resting stage. ACKNOWLEDGEMENTS
The authors thank the Brazilian National Research Council (CNPq) for its support and R.R. Chagas for her technical assistance.
REFERENCES Czekala, N.M., Hodges, J.K., Gause, G.E. and Lasley, B.L., 1980. Annual circulating testosterone levels in captive and free-ranging male armadillos (Dasypus novemcinctus). J. Reprod. Fertil., 59: 199--204. Elias, H., Henning, A. and Schwartz, D.E., 1971. Stereology: applications to biomedical research. Physiol. Rev., 51: 158--200. Enders, A.C., 1966. Reproductive cycle of nine-banded armadillo (Dasypus novemcinctus). Symp. Zool. Soc. London, 15: 295--310. Flemming, T.H., 1973. The reproductive cycles of three species of opossums and other mammals in the Panama Canal zone. J. Mammal., 54: 439--455.
141 Godinho, H.P., Cardoso, F.M., C6ser, A.M.L., Machado, A.B.M. and Nogueira, J.C., 1977. Biologia reprodutiva de machos Dasypus novemcinctus. Cienc. Cult., 2 9 : 2 7 3 (Abstr.). Hamlett, G.W.D., 1932. The reproductive cycle in the armadillo. Z. Wiss. Zool., 141: 143--157. Newman, H.H., 1913. The natural history of the nine-banded armadillo of Texas. Am. Nat., 47 : 513--539. Peppler, R.D. and Canale, J., 1980. Quantitative investigation o f the annual pattern of follicular development in the nine-banded armadillo (Dasypus novemcinctus). J. Reprod. Fertil., 59: 193--197. Peppier, R.D. and Stone, S.C., 1981. Annual pattern in plasma testosterone in the male armadillo, Dasypus novemcinctus. Anita. Reprod. Sci., 4: 49--69. Roosen-Runge, E.C. and Giesel, L.O., 1950. Quantitative studies on spermatogenesis in the albino rat. Am. J. Anat., 87 : 1--23. Sadleir, R.M.F.S., 1969. The Ecology of Reproduction in Wild and Domesticated Mammals. Methuen, London, xii + 321 pp. Snedecor, G.W. and Cochran, W.G., 1967. Statistical Methods. 6th ed. Iowa State University Press, Ames, IA, vii + 593 pp. Taber, F.W., 1945. Contribution on the life history and ecology of the nine-banded armadillo. J. Mammal., 26: 211--226. Tortes, C.N., Godinho, H.P. and Setcheil, B.P., 1981. Frequency and duration of the stages o f the cycle of the seminiferous epithelium of the nine-banded armadillo (Dasypus novemcinctus). J. Reprod. Fertil., 61: 335--340. Weaker, F.J., 1977a. The fine structure of the interstitial tissue o f the testis of the ninebanded armadillo. Anat. Rec., 187: 11--28. Weaker, F.J., 1977b. Spermatogonia and the cycle of the seminiferous epithelium in the nine-banded armadillo. Cell. Tiss. Res., 179: 97--109.
135
Elsevier Science Publishers B.V., Amsterdam -- Printed in The Netherlands
SEASONAL VARIATION IN SPERMATOGENESIS IN THE NINE-BANDED ARMADILLO (DASYPUSNOVEMCINCTUS)FROM SOUTHEASTERN BRAZIL
C.N. T O R R E S , H.P. G O D I N H O
and A.B.M. M A C H A D O
Department of Morphology, Institute of Biological Sciences, Federal University of Minas Gerais, Belo Horizonte, M G (Brazil) (Accepted 27 M a y 1983)
ABSTRACT
Tortes, C.N., Godinho, H.P. and Machado, A.B.M., 1983. Seasonal variation in spermatogenesis in the nine-banded armadilI~ (Dasypus novemeinctus) from southeastern Brazil. Anita. Reprod. Sci., 6: 135--141. Nine-banded armadillos living in the southeastern region of Brazil are seasonal breeders. The weight of the testes, diameter of seminiferous tubules, the number of type A spermatogonia and round spermatids at stage 1 of the cycle of the seminiferous epithelium, and volumetric proportions of Leydig cells showed significant variations during the annual reproductive cycle. These parameters reached a peak during the months of November and December when rainfall was plentiful and the number of daily hours of sunlight was a t its highest level. In the period of June to August the above parameters were at their lowest values.
INTRODUCTION
In a preliminary paper, Godinho et al. (1977) demonstrated a yearly variation in testis and seminal vesicle weights and in citric acid and fructose contents of the seminal vesicles of nine-banded armadillos. Czekala et al. (1980) have shown in nine-banded armadillos from the Northern hemisphere that the highest concentration of plasma testosterone occurred during the summer and that it was maintained at relatively high levels during the other seasons, indicating that testicular function continued throughout the year. They also reported that the testicularhistology of these animals was identical at all times of the year, in agreement with previous works by Weaker (1977a, 1977b), who detected no major seasonal modifications in the spermatogenic process nor in the interstitialtesticular tissue of the nine-banded armadillo. Recently, Peppler and Stone (1981) have also demonstrated that the plasma testosterone level of the same species does not significantly change during the year although it is higher in the summer than in the winter. In the present paper, histometrical analyses of testicular structures from
0378-4320/83/$03.00
© 1983 Elsevier Science Publishers B.V.
136
nine-banded armadillos living in southeastern Brazil were performed in order to evaluate the morphological changes that may occur in the testes during the annual reproductive cycle. MATERIALS AND METHODS
Thirty-one male nine-banded armadillos, 3.5--4.2 kg in body weight, were captured during the period of one year (2 animals were captured in each of the months Sep., Oct., Apr., July and Aug., and 3 in each of Nov., Dec., Jan., Feb., Mar., May and June), near Belo Horizonte (19°49 ~S). After killing the animals, their testes were removed from the abdominal cavity and then weighed (data for testes weight indicate the mean weight of right and left testes). Fragments of the testes were fixed in Susa's fluid, embedded in paraffin wax and sectioned at 10 pm. Hematoxylin-eosin staining was used throughout the study. The nuclei of type A spermatogonia, round spermatids and Sertoli cells were counted in 10 randomly selected cross-sections of seminiferous tubules at stage 1 of the cycle of the seminiferous epithelium in each animal. The diameters of these cross-sections were measured. Stage 1 encompasses the portion of the cycle extending from the shedding of the spermatozoa into the lumen of the seminiferous tubule to the beginning of spermatid nuclei elongation. This corresponds to the first stage of the cycle in the eight-stage method of Roosen-Runge and Giesel (1950) for the rat and applied to the nine-banded armadillo (Tortes et al., 1981). The single generation of spermarids present in stage 1 constitutes the group of most advanced germinal cells in the cycle; this allows determinations of their number with ease. Volumetric proportions, expressed in percentages, of testis structures were determined according to Elias et al. (1971) with the use of a 25-point ocular integrator. The coincidences between the ocular grid points and Leydig ceils, seminiferous tubule wall, tubular lumen and the other intertubular structures were recorded as obtained from 200 counts per animal. The least significant mean difference test (Snedecor and Cochran, 1967) was used to compare the monthly variations among testicular structures (P < 0.05). RESULTS
Testis weight. Testes attained their maximal weight of 8.8 +- 1.7 g in November; they decreased in weight from December until June when they reached 4.4 -+ 1.8 g (Fig. 1,F). The testes of animals killed in September and November were significantly heavier than those killed in May, June, July and August. Diameter of seminiferous tubules. The diameter of the seminiferous tubules reached the maximal value of 230 + 18.9 pm in November and decreased
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Fig. 1. Seasonal changes in testes weight (solid circles) and seminiferous tubule diameter (open circles) (F), number of spermatogonia (solid circles) and round spermatids (open circles) in stage 1 of the cycle of the seminiferous epithelium (E) and volumetric proportion, expremed in percentage, of Leydig cells (D) of nine-banded armadillos as related to daily hours of sunlight (C), temperature (B) and rainfall (A) during the period of one year near Belo Horizonte, Brazil (19°49'S); gradually to 161 + 20.3 ~m in June (Fig. 1,F). The diameter of the seminiferous tubules in November was significantly greater than those o f May, June, July and August and in December it was significantly greater than in June and August (Figs. 2,3).
Type A spermatogonia. The highest number (7.5 -+ 3.5) o f type A spermatogonia per cross~ection o f seminiferous tubules at stage 1 o f the cycle was observed in November; in the other months their number was about half of that o f November (Fig. 1,E). The number of type A spermatogonia in November was significantly greater than that of any other month.
138
Fig. 2, 3. Transverse sections of seminiferous tubules of armadillos at stage 1 of the cycle of the seminiferous epithelium (asterisks) taken at the periods of maximal (December, Fig. 2) and minimal (June, Fig. 3) spermatogenic activity in the annual reproductive cycle. H--E, 192X.
Round spermatids. The number of round spermatids (Fig. 1,E) in stage 1 of the cycle reached a peak of 259 + 65 in December. Their number gradually decreased to reach 141 + 21 in August. The number of such spermatids f o u n d in November and December was significantly greater than those of the periods o f May to June and May to August, respectively (Figs. 2,3).
139
Sertoli cells. The number of Sertoli cells did not show appreciable variation during the cycle.
Volumetric proportions of testis structures. Leydig cells were the intertubulax structures which showed the widest range of variation during the annual cycle (Fig. 1,D). They reached a peak of 11 + 1.3% in November and gradually decreased until June when they reached 2.2 + 0.7% of the testis volume. The November value was significantly greater than those of the months of February to August. The other components of the intertubular space as well as tubular lumen and tubular wall did not present marked volumetric variations during the year. Climatic parameters. Rainfall, maximal and minimal mean air temperature and hours of sunlight occurring in the region during the period of study are presented in Fig. 1 (A, B, C). DISCUSSION
An analysis on some aspects of the reproductive biology of nine-banded armadillos (Dasypus novemcinctus) of the Southern hemisphere as compared to those of the Northern hemisphere is now presented. The various histometrical parameters of the testis of the nine-banded armadillos studied in the present work showed significant annual variation (except for the Sertoli cells), in disagreement with the histological findings of Weaker (1977a, 1977b) and Czekala et al. (1980). The values for those parameters reached a peak during the months of November and December (summer in the Southern hemisphere). At this time of the year, rainfall was plentiful and the number of daily hours of sunlight was at its highest. At a corresponding season, nine-banded armadillos from the Northern hemisphere show peak testicular activity (Czekala et al., 1980; Peppler and Stone, 1981). During the period of June to August those testis parameters of the Brazilian nine-banded armadillos were at their lowest values in the annual cycle. This was the time when rainfall was almost zero and the daily hours of sunlight markedly diminished. The annual reproductive cycle of females of ninebanded armadillos has been studied mainly in specimens living in the Northern hemisphere (Newman, 1913; Hamlett, 1932; Enders, 1966; Peppler and Canale, 1980). In these animals, ovulation and fertilization occur from June to August; retarded implantation of the blastocyst takes place in November or December and is followed by a gestation period of about 4 months; birth of monozygous quadruplets occurs in February or March. The only information available on the reproductive cycle of female nine-banded armadillos in Brazil consists of unpublished data of one (ABMM) of the authors of the present paper, who found that females captured in December, in the same area as those of the present study, showed ovaries with mature follicles or corpora lutea, and the fundus of the uterus more vascularized than the rest
140
of the organ. In June, the uterus contained fetuses 1.6--2.0 cm in length; in August the fetuses measured about 8.5 cm. The young were born in September. As observed in several other wild mammals (Sadleir, 1969), a change in the reproductive cycle, associated with the inversion of the seasons of the year, is apparent in nine-banded armadillos living in the Southern hemisphere as compared with those from the Northern hemisphere. Photoperiod, temperature and rainfall are usually related to the beginning of the breeding season. Near the Equator, the very small variation in the photoperiod seems to have no effect on mammalian reproduction (Sadleir, 1969). At the latitude of the area under study, 20°S, the increase of about 1 rain day-1 in the photoperiod, between the June and December solstices, may be sufficient to affect the reproductive cycle in nine-banded armadillos. It is difficult to perceive any relationship between temperature and reproduction in our nine-banded armadillos. The maximal daily mean temperature in our region does not change considerably, remaining at around 28-32°C throughout the year, whereas the minimal mean temperature falls to around 4°C during the dry season. Over this temperature range it is possible that nine-banded armadillos may be affected only in their search for food, limiting it to the more comfortable hours of the day, as indicated by Taber (1945). There seems to be a positive correlation between minfaU and the breeding season of the nine-banded armadillo. Immediately after the first rains the availability of food increases and, as observed in mammals of Panama (Flemming, 1973), this time coincides with the weaning period. Mating would occur shortly thereafter. The available data on .the reproductive cycle of nine-banded armadillos living near Belo Horizonte suggest that these animals are seasonal breeders. Spermatogenic activity is maximal in November and December, within the rainy season, and then it progressively decreases until June, without, however, reaching a complete resting stage. ACKNOWLEDGEMENTS
The authors thank the Brazilian National Research Council (CNPq) for its support and R.R. Chagas for her technical assistance.
REFERENCES Czekala, N.M., Hodges, J.K., Gause, G.E. and Lasley, B.L., 1980. Annual circulating testosterone levels in captive and free-ranging male armadillos (Dasypus novemcinctus). J. Reprod. Fertil., 59: 199--204. Elias, H., Henning, A. and Schwartz, D.E., 1971. Stereology: applications to biomedical research. Physiol. Rev., 51: 158--200. Enders, A.C., 1966. Reproductive cycle of nine-banded armadillo (Dasypus novemcinctus). Symp. Zool. Soc. London, 15: 295--310. Flemming, T.H., 1973. The reproductive cycles of three species of opossums and other mammals in the Panama Canal zone. J. Mammal., 54: 439--455.
141 Godinho, H.P., Cardoso, F.M., C6ser, A.M.L., Machado, A.B.M. and Nogueira, J.C., 1977. Biologia reprodutiva de machos Dasypus novemcinctus. Cienc. Cult., 2 9 : 2 7 3 (Abstr.). Hamlett, G.W.D., 1932. The reproductive cycle in the armadillo. Z. Wiss. Zool., 141: 143--157. Newman, H.H., 1913. The natural history of the nine-banded armadillo of Texas. Am. Nat., 47 : 513--539. Peppler, R.D. and Canale, J., 1980. Quantitative investigation o f the annual pattern of follicular development in the nine-banded armadillo (Dasypus novemcinctus). J. Reprod. Fertil., 59: 193--197. Peppier, R.D. and Stone, S.C., 1981. Annual pattern in plasma testosterone in the male armadillo, Dasypus novemcinctus. Anita. Reprod. Sci., 4: 49--69. Roosen-Runge, E.C. and Giesel, L.O., 1950. Quantitative studies on spermatogenesis in the albino rat. Am. J. Anat., 87 : 1--23. Sadleir, R.M.F.S., 1969. The Ecology of Reproduction in Wild and Domesticated Mammals. Methuen, London, xii + 321 pp. Snedecor, G.W. and Cochran, W.G., 1967. Statistical Methods. 6th ed. Iowa State University Press, Ames, IA, vii + 593 pp. Taber, F.W., 1945. Contribution on the life history and ecology of the nine-banded armadillo. J. Mammal., 26: 211--226. Tortes, C.N., Godinho, H.P. and Setcheil, B.P., 1981. Frequency and duration of the stages o f the cycle of the seminiferous epithelium of the nine-banded armadillo (Dasypus novemcinctus). J. Reprod. Fertil., 61: 335--340. Weaker, F.J., 1977a. The fine structure of the interstitial tissue o f the testis of the ninebanded armadillo. Anat. Rec., 187: 11--28. Weaker, F.J., 1977b. Spermatogonia and the cycle of the seminiferous epithelium in the nine-banded armadillo. Cell. Tiss. Res., 179: 97--109.