36 S.-Afr. Tydskr. Dicrk. 1994,29(1) Reproductive biology of the Cape serotine bat, Eptesicus capensis in the Transvaal, South Africa

M_ van der Merwe Mammal Research Institute, University of Pretoria, Pretoria, 0002 Republic of South Africa

Received 8 FebrlUlry 1093; accepted 14 July /993

The reproductive biology of the Cape serotine bat, Eptesicus capensis, was investigated histologically. The study was based on 67 specimens collected over a six-year period. This species is seasonally monoestrou$, normally giving birth to twins during November. Spermatogenesis peaks during autumn (March-May) when masses of spermatozoa are released into the epididymides, and spermatozoa are present in the epididymides from March to October. Copulations are initiated during this period from the end of March to the beginning of April, with the first ovulations occurring during August. During the pre-ovulation period leucocytes are commonly found within the uterine lumen and uterine glands of females.

Die voortplantingsbiologie van die Kaapse dakvlermuis, Epresicus capensis, is histologies ondersoek. Die studie is gebaseer op 67 monsters wat oor In periode van ses jaar versamel is. Hierdie spesie is seisoenaal monestrus, en skenk gewoonlik geboorte aan tweelinge gedurende November. Spermatogenese bereik 'n piek gedurende die herts (Maart-Mei) wanneer massas sperme in die epididimi gestort word, en sperme is teenwoordig in die epididimi vanaf Maart tot Oktober, Kopulasie neem 'n aanvang gedurende die periode oinde Maart en begin April, terwyl die eerste ovulasies gedurende Augustus plaasvind. Gedurende die pre­ ovulasie perjode is leukosiete algemeen in die uterus lumen en ·kliere van wyfies gevind,

The Capc serotine bat Eples;cus capens;s (A. Smith, 1829) 3:1: 1:5 by volume). is a member of the family Vespcnilionidae, which is the Testes, epididymides and uteri containing embryos were largest family of the insectivorous bats (Hill & Smith 1984). prepared for histological examination. Following paraffin­ Members of this family occur throughout the world, except wax embedding all samples were serially sectioned at 5 IJ.m in polar and near polar regions (Hill & Smith 1984). and the mounted series stained with Ehrlich's haematoxylin In the Transvaal the Capc serotine bat is common and and counterstaincd with eosin.

. widespread with a wide habitat tolerance (Rautenbach )

9 1982). It is a small species with males averaging 5,9 g and Results 0

0 femals 7,3 g (Rautenhach 1982). Allhough they arc quile

2 Spcrmatogenic activity was initiated towards the end of

d common in the Transvaal, knowledge about their reproduc­ spring. Three males collected on 6 November 1987 showed e t tive biology is limited. Rautenhach (1982) collected gravid

a spcrmatogenic activity in their seminiferous tubules with d females during October and November and lactating females ( many type B spermatogonia present in some sections but no

r during September, November and December. From these e primary spermatocytes. Two males collected on 7 January h data he suggested a panurition season during the first half of s 1988 had numerous primary spermatocyles in their semini­ i l summer. The number of foetuses varies from singletons to b ferous tuhules but no secondary spermatocytes. Spermato­ u triplets (Shortridge 1914; Smithers 1971; Rautenbach 1982; genic activity was at a peak at the beginning of autumn P

e Skinner & Smithers 1990). Amongst the vespenilionids (March and April) when masses of spermatozoa were re­ h t twins arc common (Hill & Smith 1984), with triplets excep­ leased from the seminiferous tubules into the epididymides. y b tional (Stuart & Stuan 1988; Skinner & Smithers 1990). Four males collected during March and three collected

d The purpose of the present study is to investigate the

e during April had large numbers of spermatozoa in both their t reproductive biology of the Cape serotine bal. n caudae epididymides and seminiferous tubules (Table I). a r The testes of all males collected between May and Sep­ g Material and Methods e tember were spermatogenically inactive, and the seminifer­ c n A total of 67 specimens (27 males, 40 females) were col­ ous tubules were lined with Semli cells and occasional stem e c i lected from the farm Klipfontein, 30 km north-cast of Vaal­ spermatogonia only. However, large numbers of spermato­ l

r water in the Transvaal, South Africa (24'08'S /28'18'E). zoa were stored in the caudae epididymides throughout this e d Owing to small sample sizes it was necessary to combine period. Two of the three males collected on 6 November n u data from six years (1983, 1984, 1987, 1988, 1989, 1990, 1987 still had some spermatozoa in their eaudae epididy­ y

a Table I) to get a picture of the annual reproductive cycle of mides, although no sign of spermatozoa could be found in w this species. All bats were captured using two 30X6 m mist­ their seminiferous tubules. These spermatozoa were e t

a nelS (Raulenbach 1985) and were brought to the laboratory therefore left over from the previous cycle. It would G

alive where they were sacrificed using ether. Testes, epidi­ therefore appear that spermatozoa were stored in the caudae t e dymides, non-pregnant uteri and uteri with embryos were epididymides for at least six months (May-October) afler n i

b preserved in Bou.in's fluid. Uteri containing foctuses were cessation of spermatogenic activity in the testes. a

S stored in AFA (a mixture of 95% ethyl alcohol, 40% forma­ Copulations were initiated during late March and early y lin, glacial acetic acid and distilled water in a ratio of b April, and all females collected from the end of March to d e c u d o r p e R S. Afr. 1. Zoo!. 1994,29(1) 37

Table 1 Sample sizes and reproductive conditions of Table 2 Number and developmental stage of monthly samples of male and female Cape serotine conceptuses bats Embryos! Development stage of Sample size Spermatozoa in Month Female foetuses embryoslmass (g) of foetuses Leucocytes Dale Males Females Testes Epididymides Uterus in uterus Aug '88 3 Morulae 2 3 Morulae Jon '88 2 2 3 2 Young blaSltx:ym (implanting) Feb '88 0 0 4 2 Morula + young blaslOcyslS Mar '88 4 2 + + + + 5 2 Young blastocysts Apr '88 4 + + + + 6 2 Morulae '90 2 1 + + + + 1 3 Morulae May '88 t 3 + '90 3 2 + + + S", '89 t 2 0,0243 + 0,0224 Jun '90 0 0 2 2 0,0183 + 0,0\34 Jul '88 2 + + + 3 2 0,0101 + 0,0164 '90 t 2 + + + 4 3 0.0273 + 0,0271 + 0,0168 Aug '88 4 1 + 5 4 0,0109 + 0,0113 + 0,0107 + 0,0013 Sep '89 5 5 + Oct'S3 2 O,t245 +0,1055 Oct '83 0 5 2 2 O,It64 + 0,0918 2 '84 0 3 2 (ncar term) 0,9800 + 0,8211 Nov '83 0 3 4 2 (fuU term) 1,6494 + 1,4816 '81 3 0 +

Spermatozoa: - = absent, + = presenl . Leucocytes: - = absen!, + = present. 1980) were orientated with their heads towards the epithelial lining, with many resting against it. The spermalOzoa in the intramural part of the UTJ were, as in all the other females examined, not orientated in any specific manner. Ovulation occurred during the second half of August. All four females collected during July (29th 1988 and 31st 1990; Table 1) had mature Graafian follicles in their ovaries.

. However, all seven females collected during August (24th )

9 1990; Table 2) had small embryos at various stages of 0

0 development. The majority were morulae in the uterine 2 lumen, with the oldest one being a young implanting blasto­ d e t cyst (Table 2). Five females collected during September all a d had small foctuses at the limb bud stage (Table 2). (

r Most gravid uteri contained two conceptuses (one in each e h hom), although singlclOns, triplets and even quadruplets s i l 20)lm were noted. b u Four of five females collected at the end of October had P

e Figure I LeucocyIcs (arrows) in the uterine lumen of a female large foctuses while one had already given birth and was h t

Cape seroline bat Collected during May 1988. lactating. One of the gravid females had two full-term y

b foclUses that were fully pigmented and near birth (Table 2).

d The combined mass of the two foctuses was 3,13 g. e t late July had some spermatozoa in the proximal ends of

n comprising 43% of the average body mass of non-gravid

a their uterine horns (Table 1). During the same period all the r females. Three females collected at the end of November g

females examined also had polymorphonuclear leucocytes in

e were all lactating. Since parturition probably occurred from c their uterine lumina and uterine glands (Figure 1, Table 1). n the end of October 10 mid-November and the majority of e No leucocytes, however, were found in the uterotubal c

i ovulations occurred during the second half of August, the l

junction area (UTJ) or oviducts. During Mareh and April, r gestation period is estimated to be about 12 weeks. e when copulations began, leucocyte numbers were low d

n compared 10 May when large numbers of leucocytes were u Discussion present Females collected at the end of July had relatively y a few leucocytes in their uterine horns. The majority of southern African insectivorous bats so far w e Spermatozoa found in the uterine lumen of all females investigated are monoestrous with one seasonal restrous t a examined were not orientated in any specific manner, and cycle per annum (sec Rautenbach 1982; Skinner & Smithers G

t showed no relationship with the epithelial lining of the 1990 for reviews). This appears to be the norm for most e

n uterine horns. No specific orientation of the spermatozoa insectivorous bats (Wimsatt 1975; Hill & Smith 1984). i b could be found in the UTJ either, with the exception of one However, recent evidence indicates that some South African a S

female collected during May in which the spermatozoa in bats deviate from this pattern. The little free-tailed bat, y

b the colliculus tubaricus part of the UTJ (see Mori & Uchida Tadarida pumiia, is polyocstrouS with up to three pregnan- d e c u d o r p e R 38 S.-Afr. Tydskr. Dierk. 1994. 29(t)

cies per breeding cycle (Van der Merwe, Rautenbach & Van which pan of the tract fulfils this function, but that useful der CoIf 1986). This is facilitated by a post-partum oestrus criteria are the identification of special relations, such as the (Van der Merwe, Giddings & Rautenbach 1987). RecenUy it attachment between spermatozoa and their storage organs or has been shown that twins are common amongst certain the presence of healthy spermatozoa. Although spermatozoa southern African bat species, and occasionally even triplets in the UTJ of the majority of Cape serotine bats examined have been documented. viz. the yellow house bat ScO/ophi­ were not orientated in any specific manner, no Icucocytcs Ius dinganii (Rautenbach 1982), the lesser yellow house bat were noted in this area, and it would therefore appear that ScolOphilus borbonicus (Van der Merwe, Rautenbach & this is the area where spermatozoa are present at least for Penzhorn 1988), and Schlieffen's bat Nycliceius schlieffenii some period of time. Whether spermatozoa in the UTJ are (Van der Merwe & Rautenbach 1986). Although twins are stored there from the onset of copulations (late March/early common in the Cape serotine bat, singletons, triplets and April) until fertiliz3tion (August) is doubtful. The fact that even quadruplets (Lynch 1989; Van der Merwe 1990; pre­ specific orientation of spermatozoa in the UTJ was observed sent study) have been recorded for this species. in only one female, as well as the leucocytic activity found Vespertilionid and rhinolophid bats inhabiting temperate in the uterine lumina of the majority of females, even as late latitudes undergo spermatogenesis during summer, and ma­ as 3 I July, place a shadow of doubt on the females ability to ting is initiated during autumn (Racey 1982). Spermatozoa store spermatozoa for prolonged periods. It is expected that are stored in the female reproductive tract and retain their leucocytes should be capable of removing all spermatozoa fertilizing eapacity until spring, when ovulation occurs and from the initial copulations (late Mareh/early April) within the females become pregnant (Wimsatt 1969; Racey 1979). the four-month period early April to the end of July. This Male bats also store spermatozoa for prolonged periods, argument is supported by the fact that the volume of even after the females have ovulated (Racey 1973, 1979). It spermatoz.oa found in the proximal parts of the uterine horns seems that the responsibility for sperm storage is shared of this species, at any stage of the study, was absolutely between the sexes, with different sexes taking greater or minute when compared with that found in the coexisting lesser shares, depending on the species (Racey 1982). Racey rusty bat. In the latter species, the uterus is massively (1979) mentioned that spermatozoa can be stored in the distended with semen (Van der Merwe & Rautenbach 1990). female tract for up to seven months, and may remain in the Therefore, the presence of spermalOzoa and leucocytes in caudae epididymides of males for up to ten months. In the uterine lumina of females collected even as late as 31 South Africa sperm storage has also been recorded in July would rather suggest more recent copulations with vespertilionids (Bernard 1982; Van der Merwe & Rauten­ rresh leucocyte activity. Funhcnnorc, as no conspicuous bach 1987, 1990) and in rhinolophids (Bernard 1983). decrease in sperm volume was noted in the uterine lumina In the Cape serotine bat the testes become completely as the winter progressed, it would rather appc3r that the . ) involuted and inactive during May, with spermatozoa being burden of prolonged sperm stomge in this species falls on 9

0 stored in the caudae epididymides at least until October. In the male, and that periodic winter copulations combined 0 2

this species spermatozoa are still present in the caudae with leucocytic aClivity, prevented a significant increase or d

e epididymides for about two months after fertiliz3tion has decrease in sperm volume. 1n the Northern Hemisphere t a occurred. In the northern Transvaal bushveld. Cape serotine copulations during winter are well-documented in bats d (

bats are frequenUy active during winter and do not appear to (Gilbert & Stebbings 1958; Wimsatt 1945). In South Africa r e go into prolonged periods of torpor, as evidenced by the fact (Transvaal Province), Van der Merwe & Rautenbach (1987) h s i that bats were taken during winter. This is not a strange argued that there is a possibility that subsequent matings l b phenomenon, as bat activity during winter has been recorded during winter may occur in the vcspcrtilionid Nycliceius u P

for much colder climates. In England, Ransome (1968, schlieffenii. However, in the vcspertilionid Pipislrellus e

h 1971) found that the greater horseshoe bat Rhinolophu., rusticus, which coexists with the Cape serotine bat, the t

y ferrwnequinum arouses to feed regularly during the winter absence of leucocyte activity and frequent copulations b hibernation period, and Racey (1982) mentioned that during winter are the result of a three-fold increase in sperm d e

t although there is evidence that repeated arousal and frequent volume between April and July (Van der Merwe & n activity during hibernation may be inimical to the storage of Rautenbach 1990). a r g spermatozoa, it is clear that bats are often naturally active At present detailed knowledge regarding reproduction in e

c for a considerable part of the period of sperm storage. South African vespertilionids is restricted to only a few n

e In most mammals, including bats. there is a massive specics. Nevertheless, in these few species three different c i l accumulation of leucocytes in the uterus shortly after reproductive delay phenomena have been found, which to a r

e insemination occurs (Hill & Smith 1984). However, in those greater or lesser extent overlap with the winter months. 1n d

n bat species that store spermatozoa in the female tract, the Schreiber's long-fingered bat Miniopterus schreibersii u reproductive tracts are remarkably free of leucocytes (Racey (Bernard 1980; Van der Merwe 1980) and the lesser long­ y a 1979) and this onslaught does not occur (Hill & Smith fingered bat M. fralerculus (Bernard 1980) delayed implan- w e 1984). The presence of both spermatozoa and leucocytes in 13tion oecurs, lasting approximately four months in t a the uterine lumina and glands of female Cape serotine bats Schreiber's long-fingered bat, both in the Natal Midlands G

t collected at different times during the winter suggests that (Bernard 1980) and the Transvaal Highveld (Van der Merwe e

n spermatozoa are not stored in the uterine lumen. The sites of 1980). i b sperm storage in female bats vary with species and can be Sperm storage or delayed ovulation has been described a

S either the oviduct, UTJ or the uterus (see Racey 1979). N. &

for Schlieffen's bat schlieffenii (Van der Merwe y

b According to Racey (1979) it is not always easy to decide Rautenbach 1987), the rusty bat P. rusticus (Van der Merwe d e c u d o r p e R S. Afr. J. Zool. 1994.29(1) 39

& Rautenbach 1990) and the Temminck's hairy bat Myotis RACEY. P.A. 1982. Ecology of bal reproduction. In: Ecology of tricolor (Bernard 1982). A third delay phenomenon de­ bats. (ed.) T.H. Kunz. Plenum Press, New York. scribed for the lesser yellow house bat Scotophilus borboni­ RANSOME. R.D. 1968. The distribution of the Greater horse· cus is retarded embryonic development after implantation shoe bat. Rhinolophus /errumeqwinum. during hibernation. in (Van der Merwe et al. 1988). relation to environmental factors. 1. 7.nol., Lond. 154: 77-112. Both Schlieffen's bat and the lesser yellow house bat RANSOME, R,O. 1971. The effect of ambient'temperatureon coexist at ca. 29"S in the northern Kruger National Park. [n the arousal frequency of the hibernating Greater horseshoe both species a reproductive delay of approximately three bat. Rhin.olophus/errumequinum, in relation to site selection months occurs. In the lesser yellow house bat the delay is and hibernation !'tate. J. 7AJol., Lond. 164: 353-371. caused by retarded embryonic development which occurs RAUTENBACH.I.L. 1982. Mammals of lIle Transvaal. Ecoplan Monograph No. I, Pretoria. directly after implantation and lasts from ApriVcarly May to RALn'ENBACH. I.L. 1985. A new technique for the efficient use the end of July (Van der Merwe et al. 1988). [n Schlieffen's of macro-mistnclS. Koedoe 28: 81--86. bat the delay is caused by the storage of spermatozoa, in SHORTRItx.E. G.c. 1934. The mammals of Soulll West Africa. both sexes, from June to the end August (Van der Merwe Vols. I & n. Heineman, London. 1987). In both species the young arc born during November. SKINNER, J.D. & SMITHERS, R.H.N. 1990. The Mammals of Despite differences during the delay period, such as the Southern African Subregion. University of Pretoria. sperm volume, sperm storage sites and the presence of Pretoria. leucocytes, the reproductive cycles of the rusty bat (Van der SMITHERS. R.H.N. 1971. The mammals of Rotswana. Museum Merwe & Rautenbach 1990) and the Cape serotine bat (pre­ Memoir No, 4. Trustees of the National Museums of sent study) coexisting in the northern Tmnsvaal bushveld at Rhodesia. Salisbury. ca. 23"S are very similar. [n both species spermatozoa arc STUART. C. & STUART. T. 1988. Veldgids 101 die Soogdiere present simultaneously in the caudae epididymides of the van Suider-Afrika. Struik. Kaapstad. male and genital LIact of the female for approximately five VAN DER MERWE. M. 1980. De]ayed implantation in the Natal months of the year (April-August) before ovulations and clinging bat Miniopterus schreibersii natalensis (A. Smith. 1834). Proc. 5th Int. Bat Res. Conf.. (cds) D.E. Wilson & fertihation occur. [n both species the majority of births A.L. Gamer. pp. 113-123. Texas Tech Press. Lubbock. Texas. occur during November. U.S.A. Acknowledgements VAN DER MER WE. M. 1990. Polytocy in the Cape serotine bat Epte.ficus capen.fi.f (A. Smith 1829) from the southern African Thanks are due to Dr I.L. Rautenbach of the Transvaal subregion. S. Afr. f. 7.001. 25: 200-203. Museun for kindly allowing me the use of his equipment for VAN DER MERWE. M .. GIDDINGS, S.R. & RAlJfENBACH,

. the capturing of specImens, as well as Mr C. Baber for ) I.L. 1987. Post-partum oestrus in the little free-lailed baL. 9 allowing me free access to localities on his farm. A special 0 Tadarida (Chaerephon) pumila (Michrochiroptcra: 0 word of thanks to Mr H.O. Niemandt for assisting me in the 2 Molossidae) at 24'S. 1. Zool .. Lond. 213: 317-326.

d histology labomtory. VAN DER MER WE. M. & RAlJfENRACH, I.L. 1986. Multiple e t

a births in Schlicffen's bat. Nycliceius schl~ffe"ii (Peters, 1859) d

( References (Chiroptera: Vcspcrtilionidae) from the southern African r

e BERNARD. R.T.F. 1980. Reproductive cycJcs of Miniop1erus subregion. S. A/r. 1. l.LJol. 21: 48-50. h s schreibersi nalaiensiJ (Kuhl, J 819) and MinioplerU"f; VAN DER MER WE. M. & RAUTENRACH.I.L. 1987. i l

b jratercuiusThomas and Schwann, 1906. Ann. Transv. Mus. Reproduction in Schlicffcn's bal, Nycliceius .f(:hJieffe"ii, in the u 32: 55-64. eastern Transvaallowveld. South Africa. 1. Repmd. Fed. 81: P

e BERNARD. R.T.F. 1982. Monlhly changes in Ihe female 41-50. h t reproductive organs and the reproductive cycle of MYOlis VAN DER MER WE. M. & RAUTENBACH. [.L. 1990. y lricolor (Vespcrtilionidac: Chiroptera). S. II/r. J. 7.001. 17: b Reproduction in the Rusty bat. Pipistrellus rusticu.f, in the

d 79--1!4. northern Transvaal bushveld. South Africa. 1. Repmd, Fert. e t BERNARD, R.T.F. 1983, Reproduction of Rhin%phus clivosus 89: 537-542. n

a (Microchiroptcra) in Natal, South Africa. Z. SdugelierkuNie

r VAN DER MER WE. M .. RAUTENBACH, I.L. & VAN DER g 48: 321-329. COLF. W.J. 1986. Reproduction in females of the little free­ e

c GILBERT. P. & STEBBINGS. R.E. 195R. Wintcr roosts of bats tailed bat. Tadarida (Chaerephon) pumila. in the eastern n

e in West Suffolk. Proc. zool. Soc. LoN!. 131: 329-333.

c Transvaal. South Africa. 1. Reprod. Ferl. 77: 355-364, i

l HILL. J.E. & SMITH. J.D. 1984. BalS: a Nalural History. VAN DER MER WE, N .. RAUTENBACH, I.L. & PENZHORN, r University of Tcxas Press. Austin. e B.L. 1988. A new pattern of early embryonic development in d LYNCH. CO. 1989. The Mammals of the north-ea... tern Cape n the sea... onally breeding non-hibernating lesser yellow house u Province. Mem. MS. Mus .. Bloemfontein 25: 1-116.

y baL. Scolophilus borbonicus (E. Geoffroy, 1803) (Oiiroptera; MORI. T. & UCHIDA. T.A. 1980. Sperm Slorage in Ihe a Vespcrtilionidae).Ann. Trans\!. Mus. 34: 551-556. w reproductive tract of the female Japanese long-fingered bat, e t Miniopterus schreibersiifuliginosus. 1. Reprod. Ferl. 58: WIMSATI, W.A. 1945. Notes on breeding behaviour. a 429-433. pregnancy, and parturition in some vespertilionid bats in the G t RACEY. P.A. 1973. The Viability of spermatozoa after prolonged eastern United States. 1. Mammal. 26: 23-33. e n storage by male and female European bal". Period. BioI. 75: WIMSA TI. W.A. ]969. Some interrelations of reproduction and i b 201-205. hiremation in mammals. Symp. Soc. Exp, Bioi. 23: 511-549. a S

RACEY. P.A. 1979. The prolonged storage and survival of WIMSATI, W.A. 1975. Some comparative aspects of y

b spcrmato7.0a in Chiroptera. J. Reprod. Ferl. 56: 391-402. implantation. Bioi. Reprod. 12: 1-40. d e c u d o r p e R