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Pollination Biology of Piperaceae Species in Southeastern Brazil
Annals of Botany 85: 455±460, 2000 doi:10.1006/anbo.1999.1087, available online at http://www.idealibrary.com on
Pollination Biology of Piperaceae Species in Southeastern Brazil RO D O L FO A N T OÃ N I O D E F I G U E I R E D O *{ and MA R L I E S S A Z I M A{ {Departamento de CieÃncias, Faculdade de CieÃncias e Letras Padre Anchieta, Rua Bom Jesus de Pirapora 140, 13207-660 JundiaõÂ, SaÄo Paulo, Brazil and {Departamento de BotaÃnica, Universidade Estadual de Campinas, 13083-970 Campinas, SaÄo Paulo, Brazil Received: 19 August 1999 Returned for revision: 22 October 1999 Accepted: 2 December 1999 Phenology, ¯oral biology and breeding system are described for 14 Piperaceae species at two sites covered by semideciduous forest in southeastern Brazil. Flowering occurred continuously throughout the year, with a peak during the windy months. All species had minute odorous and nectarless ¯owers visited by insects; ®ve of them showed a substantial degree of self-compatibility, and one was andromonoecious. Seven species were wind pollinated, and three were exclusively pollinated by insects. Flower visitors were mainly hover¯ies and bees. # 2000 Annals of Botany Company Key words: Piper spp., Ottonia spp., Pothomorphe umbellata, pepper family, phenology, ¯oral biology, breeding system, ambophily, semi-deciduous forest.
I N T RO D U C T I O N The family Piperaceae is pantropical with species distributed in the New World from Mexico to southeastern Argentina. Brazilian forests harbour 283 species of Piper L., 25 species of Ottonia Sprengel, and two species of Pothomorphe Miquel (Yuncker, 1972, 1973, 1975). In spite of the importance of the pepper family in tropical forests, the biology of its species is almost unknown in Brazil. Until recently little was known about the pollination biology of the paleotropical Piper nigrum L. (black pepper), the most well known of the Piperaceae due to its economic importance (Pouvreau, 1984; Richards, 1997). Martin and Gregory (1962) recorded wind as the most important pollen vector in black pepper, while pollination of neotropical Piper species is thought to be carried out by bees, beetles and drosophilid ¯ies (Janzen, 1991; McDade et al., 1994; Kricher, 1997), mainly on account of the studies made in Central America (Semple, 1974; Fleming, 1985). In South America, there have been no studies on the pollination biology of Piperaceae species. The present study describes ¯oral morphology and the breeding and pollination systems of 11 species of Piper, two of Ottonia, and one of Pothomorphe in a semi-deciduous forest of southeastern Brazil. Pollination biology is discussed in the light of what is known for black pepper and the Central American Piper species. M AT E R I A L S A N D M E T H O D S Study area and plant species Fieldwork was undertaken from 1 Mar. 1994 to 31 Dec. 1995 at the Reserva de Santa Genebra (228490 S, 478070 W, * For correspondence. E-mail roro®[email protected]
0305-7364/00/040455+06 $35.00/00
670 m alt.) and at the EstacËaÄo Experimental de JundiaõÂ /IAC (238060 S, 468550 W, 715 m alt.), two reserves of semideciduous forest in southeastern Brazil ( for geographical and vegetation descriptions see Morellato, 1992; Morellato and LeitaÄo-Filho, 1995). There is a marked seasonality in the study area, with a dry, cold season from April to September and a wet, warm season from October to March (Mello et al., 1994). The wind velocity increases at the end of the dry season (September). Piperaceae species were surveyed by transects made in the dry and swampy parts of the reserves. Species names follow Yuncker (1972, 1973), and voucher specimens were deposited at the herbaria of Universidade Estadual de Campinas (UEC) and Instituto AgronoÃmico de Campinas (IAC) (see Appendix). Phenology The phenology was registered through weekly observations on ®ve individuals of each Piperaceae species, from 1 Jan. to 31 Dec. 1995. The relationship between ¯owering periods and climate variables was tested using Spearman, correlation coecient, rs (Zar, 1996). Daily observations were carried out on in¯orescences to verify the period of anthesis and the temporal separation of stigma receptivity and anther dehiscence (types of dichogamy following Lloyd and Webb, 1986). Flower characteristics Flower morphology was described using ten in¯orescences of each species. Names of ¯oral parts follow Radford et al. (1974) and Weberling (1992). Colour names follow Smithe (1975). Stigma receptivity was checked with the hydrogen peroxidase reaction (Zeisler, 1938). # 2000 Annals of Botany Company
De Figueiredo and SazimaÐPollination in Piperaceae
Osmophores and nectaries were tested with neutral red 1 : 1000 for odour (Dafni, 1992) and 1 : 10 000 for nectar (Kearns and Inouye, 1993). Breeding system In order to analyse the occurrence of within-in¯orescence fertilization (autogamy and geitonogamy), 30 immature in¯orescences were isolated in waterproof bags, excluding pollen from other plants. Thirty unbagged, randomly chosen in¯orescences of the same age were left untreated as controls. The number of fruits that set and survived until day 20 after the ¯owering period was checked and used as an index of successful pollination. Pollination system Wind pollination experiments were conducted by isolating 30 immature in¯orescences in thin cloth bags that completely excluded visitors. Thirty unbagged in¯orescences of the same age were observed as controls. Fruit set was checked 20 d after ¯owering and used as an index of successful pollination. Open ¯owers of each Piperaceae species were observed for 60 h, from 0600 to 1800 h for veri®cation of insect visitors. The insects collected were identi®ed and deposited at the Identi®cation Centre of Phytophagous Insects (CIIF) of Universidade Federal do ParanaÂ. The behaviour of insect species was monitored by ®eld observations and the presence of pollen on their body parts was checked with a stereoscopic microscope. R E S U LT S A N D D I S C U S S I O N The plants and their phenology
Flower characteristics The Piperaceae in¯orescences show a variable number of minute ¯owers (Table 1, Fig. 2A), which are creamy, yellowish or whitish in colour (Table 1). These colours are also the most common for the Piperaceae in Central America (Tebbs 1989a, b), but Tebbs found some Piper species with pink and violet in¯orescences. Almost all species produce a sweet/lemon-lime odour, except for P. aduncum whose ¯owers are scentless. Lemon-lime odour occurs in some Mesoamerican Piperaceae species (Tebbs, 1989a). This odour is probably the most widespread in the family. Nectar is not discernible in the ¯owers, as has been observed for other Piper species (Fleming, 1985; Marquis, 1988; Ollerton, 1996). The ¯owers are astylocarpellous in Piper gaudichaudianum, P. glabratum, P. macedoi and Pothomorphe umbellata, astylocarpepodic in Ottonia species, and stylocarpellous in the other Piperaceae species. The style is absent in Piper gaudichaudianum, P. glabratum and P. macedoi, as well as in the Ottonia and Pothomorphe species. All other species are homostylous. The stigmas are decurrent in almost all species, except for P. amalago and P. arboreum, whose stigmas are clavate. Piper and Pothomorphe species commonly show three stigmas, while Ottonia species show four stigmas. The stamens are ®lantherous, their number varying from three to six in the species studied (Table 1). The stamens of Piper arboreum number four per ¯ower, but Tebbs (1989a) found six stamens in Central American populations. All Piperaceae species have divergent stamen thecae, with extrorse longitudinal dehiscence of the anthers, which drop after the pollination period. Pollen of all species is dry and of the monad type, white or yellow coloured.
12
45
10
40 35
8
30 25
6
20
4
15
Wind velocity
Most of the Piperaceae studied are shrubs, with the exception of Piper arboreum, which is a treelet, and P. mikanianum, which has an herbaceous habit. Piper gaudichaudianum and Ottonia martiana are found only in swamps, while the other species grow both in and at the edge of swamps and in dry parts of the semi-deciduous forest. The phenological pattern of the studied Piperaceae is of the continual type (sensu Newstron et al., 1994). The peak of ¯owering species coincides with the peak of wind velocity in the area (Fig. 1), although a strong correlation was not veri®ed (rs 0.56, 0.05 5 P 5 0.10). Some climatic variables have been correlated with the ¯owering periods of Central American Piper species (Opler et al., 1980; Marquis, 1988; Wright, 1991), but such relations were not observed in the species studied here. The proximal ¯owers in the in¯orescences open ®rst in the majority of species, with the exception of Piper mikanianum, P. xylosteoides and Pothomorphe umbellata, whose ¯owers all open at the same time. Stigma receptivity, as well as anther dehiscence, last up to 6 d in all species. In¯orescences are in bloom for up to 10 d, while Marquis (1988) found a ¯owering time of 1 week for the Mesoamerican species Piper arieianum C.DC.
The majority of the studied species exhibits incomplete protogyny, but Piper mikanianum shows complete protogyny, P. xylosteoides incomplete protandry, and P. regnelli homogamy. The primary type of dichogamy found in Piperaceae is incomplete protogyny, which has also been recorded for paleotropical species (Menon, 1949; Martin and Gregory, 1962).
No. of species flowering
456
10
2
5
0
0 J
F M A M J
J
A
S
O N D
Months FIG. 1. Number of Piperaceae species ¯owering (bars) and wind velocity (line) in semi-deciduous forest of southeastern Brazil in 1995.
De Figueiredo and SazimaÐPollination in Piperaceae
457
T A B L E 1. In¯orescence and ¯ower characteristics of Piperaceae species in semi-deciduous forest of southeastern Brazil Species Piper aduncum P. amalago P. arboreum P. crassinervium P. gaudichaudianum P. glabratum P. macedoi P. mikanianum P. mollicomum P. regnelli P. xylosteoides Ottonia martiana O. propinqua Pothomorphe umbellata
Number of ¯owers (x + s.d.) 505.00 + 19.13 351.40 + 28.50 403.60 + 18.23 544.00 + 18.51 407.20 + 14.54 446.00 + 53.20 607.20 + 19.54 199.20 + 8.29 577.20 + 54.21 746.20 + 40.06 290.20 + 33.12 118.80 + 2.59 284.60 + 18.05 449.00 + 39.40
Colour
Odour intensity
Length of the style (mm)
Length of the ®lament (mm)
Stamen number
±
0.5 1.5 1.0 2.0 sessile sessile sessile 0.5 0.5 1.0 0.2 sessile sessile sessile
0.5 3.0 4.0 2.4 0.3 0.5 0.5 1.0 1.0 1.2 0.2 0.8 1.0 ±
6 4 4 4 3 4 4 3 3 3 3 4 4 2
Cream Pale horn Sulphur yellow Spectrum yellow Sulphur yellow Pale horn Sulphur yellow Sulphur yellow Pale horn Straw yellow Cream diluted Sulphur yellow Sulphur yellow Pale horn
Some of these androecium features have previously been described for Piper (Yuncker, 1972, 1973; Endress, 1994). Breeding systems Most of the Piperaceae species are hermaphrodite, with the exception of Piper arboreum which has some in¯orescences with only staminate ¯owers and others with hermaphrodite ones. Andromonoecy has not previously been recorded in Piperaceae (see Tebbs, 1993, for a review of the family characteristics), although Ollerton (1996) noticed an individual of a monoecious Australian Piper species bearing only male in¯orescences. The ratio of staminate to hermaphrodite in¯orescences is 0.53 in P. arboreum, which shows the lowest fruit formation amongst the Piperaceae (Table 2). Five species show a substantial degree of selfcompatibility: Piper aduncum, P. mollicomum, P. regnelli, P. gaudichaudianum and Pothomorphe umbellata. Piper macedoi and P. xylosteoides set few fruits, suggesting that they manifest an unimportant degree of self-compatibility or that the bag initially contained few very small insects (such as thrips) carrying pollen from another plant. The other Piperaceae species are necessarily outcrossed (Table 2). As the microclimate inside the waterproof bags may have been very dierent to that outside (see Corbet and Willmer, 1981), and minute insects such as thrips may have been present inside them, the estimation of self-pollinated fruit development (Table 2) is conservative. The breeding systems of Piperaceae are known for only very few species: the paleotropical Piper nigrum and the neotropical P. arieianum which are self-compatible (Martin and Gregory, 1962; Marquis, 1988; Sasikumar et al., 1992), and the paleotropical P. methysticum which is selfincompatible (Prakash et al., 1994). Although tests for agamospermy were not carried out, Piper aduncum and Pothomorphe umbellata may be agamospermous, since all in¯orescences of these species developed fruits. It has been suggested that agamospermy occurs in Piper nigrum (Gentry, 1955).
Pollination systems Wind pollination occurs in seven of the studied Piperaceae: Piper arboreum, P. gaudichaudianum, P. macedoi, P. mikanianum, P. xylosteoides, Ottonia martiana and O. propinqua (Table 2). Anemophily has previously been recorded only for P. nigrum in Panama (Martin and Gregory, 1962), whereas other Central American Piper species are not wind pollinated (Semple, 1974; Fleming, 1985). As the bags used for insect exclusion may have aected access by windblown pollen, the estimation of wind pollination (Table 2) may be conservative. Furthermore, small insects initially present in the bag may have also contributed to fruit set by autogamy or geitonogamy, as perhaps occurred in P. crassinervium. Four species, P. aduncum, P. mollicomum, P. regnelli and Pothomorphe umbellata, did not yield substantially more fruits in insectproof bags than could have been accounted for by withinin¯orescence fertilization in waterproof bags, hence they were not considered to be wind pollinated. The majority of species grow at the edge of the forest, an appropriate place for pollen transport by wind (Arruda and Sazima, 1988), but Piper arboreum, P. gaudichaudianum, P. mikanianum, P. regnelli, O. martiana and O. propinqua also grow in the forest under-storey. These individuals, too, are eectively pollinated by wind (Table 2). Bawa and Crisp (1980) and Bawa et al. (1985) also recorded some anemophilous under-storey species within the Moraceae. Species of Piperaceae exhibit some classic traits of anemophilous plants, such as the increase of individuals ¯owering in windy months ( from September to December, see Fig. 1), the small and pulverulent pollen grains and the protogynous ¯owers (Lloyd and Webb, 1986; Honig et al., 1992). Piper amalago, P. crassinervium and P. glabratum may be exclusively insect-pollinated. As insects visit in¯orescences of all Piperaceae species (Table 2), these plants may well bene®t from insect pollination, although we have no evidence of this since none of our treatments excluded wind pollination. Flies (mainly hover¯ies, Syrphidae: Diptera) and bees (mainly Apidae: Hymenoptera) were the most frequent visitors, transporting Piperaceae pollen
458
De Figueiredo and SazimaÐPollination in Piperaceae
FIG. 2. A, In¯orescence of Ottonia martiana showing its minute ¯owers. A small hover¯y, Ocyptamus sp. (Diptera: Syrphidae), length approx. 8 mm, is visiting the in¯orescence. B, Copestylum tripunctatum (Syrphidae), 15 mm in length, visiting a hermaphrodite in¯orescence of Piper arboreum.
on their bodies (Fig. 2B). Central American Piper species are pollinated mainly by stingless bees of the genus Trigona (Apidae: Hymenoptera) (Semple, 1974; Fleming, 1985). Hover¯ies are common and generalist ¯ower visitors, being attracted by creamy-coloured and scented ¯owers,
even of anemophilous plant species (Robertson, 1928; van der Goot and Grabrandt, 1970; Stelleman and Meeuse, 1976; Waldbauer, 1984; Barth, 1991; Arruda and Sazima, 1996). Hover¯ies and bees visit Piperaceae ¯owers for pollen. Moving along the in¯orescence, the ¯ies touch the
De Figueiredo and SazimaÐPollination in Piperaceae
459
T A B L E 2. Number of Piperaceae in¯orescences pollinated by sel®ng, wind and open pollination (control), as well as receiving insect visitors, in semi-deciduous forest of southeastern Brazil Within-in¯orescence fertilization
Open pollination (control)
Wind pollination
Open pollination (control)
30
28
30
30
P. amalago
0
22
0
24
P. arboreum
0
6
8
7
P. crassinervium
0
15
1
13
P. gaudichaudianum
6
20
23
19
P. glabratum
0
15
0
18
P. macedoi
2
18
15
15
P. mikanianum
0
14
12
12
P. mollicomum
17
22
14
23
P. regnelli
30
27
23
28
P. xylosteoides
2
6
12
8
Ottonia martiana
0
24
27
25
O. propinqua
0
30
30
30
16
16
12
12
Species Piper aduncum
Pothomorphe umbellata
open anthers with the labell and gather pollen with the pulvilli and arolia. In some instances, hover¯ies also touch the bracts with the labell, presumably collecting pollen grains or some other substance. The bees move faster along the in¯orescence, collecting pollen with their legs. Flies visit several in¯orescences before leaving the plant, whereas bees visit only two to four in¯orescences, both of them moving from the proximal part of the in¯orescence to the distal part. The behaviour of these visitors may increase autogamy and geitonogamy (Barth, 1991; Richards, 1997). Species of Coleoptera and Hemiptera were also found in Piperaceae in¯orescences, but they rarely moved between ¯owers and behaved as herbivores, chewing and sucking ¯owers and fruits. Thomisidae spiders prey upon bees and ¯ies visiting the in¯orescences of Piper crassinervium and P. regnelli.
CO N C L U S I O N S This study has identi®ed ambophily (wind and small insects) as the primary pollination system for southeast Brazilian Piperaceae species. This is a well-based hypothesis, since all Piperaceae species show attributes commonly associated with anemophily and entomophily, but one which had not
Visiting insects
Number of visits (%)
Diptera Hymenoptera Lepidoptera Diptera Hymenoptera Diptera Hymenoptera Diptera Hymenoptera Lepidoptera Diptera Hymenoptera Diptera Hymenoptera Lepidoptera Diptera Hymenoptera Diptera Hymenoptera Diptera Hymenoptera Diptera Hymenoptera Lepidoptera Diptera Hymenoptera Diptera Hymenoptera Diptera Hymenoptera Lepidoptera Diptera
255 8 1 241 56 165 36 663 41 5 16 5 131 15 1 75 10 95 17 103 8 1500 604 2 47 9 35 2 81 106 1 8
(96.59) (3.03) (0.38) (81.21) (18.79) (82.09) (17.92) (92.86) (5.78) (0.71) (76.19) (23.81) (89.12) (10.20) (0.68) (88.24) (11.76) (84.82) (15.18) (92.79) (7.21) (71.23) (28.68) (0.09) (87.04) (16.07) (94.74) (5.26) (43.09) (56.38) (0.53) (100.00)
previously been demonstrated experimentally. Besides the continual ¯owering phenology, the incomplete protogyny, the odorous in¯orescences and the self-pollination, the study detected an andromonoecious species, the ®rst record of andromonoecy for this plant family.
AC K N OW L E D G E M E N T S The authors thank Conselho Nacional de Desenvolvimento Cientõ ®co e TecnoloÂgico (CNPq, process number 140055/ 94-0) for ®nancial support. This work was presented at the XVI International Botanical Congress (Saint Louis, USA) and R. A. de Figueiredo also acknowledges fellowships from FundacËaÄo de Amparo aÁ Pesquisa do Estado de SaÄo Paulo (FAPESP, process number 1999/04469-8) and from XVI IBC Fellowship Committee. Insect identi®cations were carried out by L. Marinoni, Pe. J. S. Moure, C. Carvalho, L. M. Almeida, O. Mielke and K. Zanol. FundacËaÄo Jose Pedro de Oliveira and EstacËaÄo Experimental de Jundiaõ / IAC provided permissions to work on their forest reserves. We are indebted to S. Corbet and D. Inouye for providing valuable comments and suggestions that improved an earlier version of the paper.
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Opler PA, Frankie GW, Baker HG. 1980. Comparative phenological studies of shrubs and treelets in wet and dry forests in the lowlands of Costa Rica. Journal of Ecology 68: 167±186. Pouvreau A. 1984. Les plantes aÁ eÂpices. In: Pesson P, Louveau J, eds. Pollinisation et productions veÂgeÂtales. Paris: INRA. Prakash N, Brown JF, Yue-Hua W. 1994. An embryological study of Kava, Piper methysticum. Australian Journal of Botany 42: 231±237. Radford AE, Dickison WC, Massey JR, Bell CR. 1974. Vascular plant systematics. New York: Harper & Row Publishers. Richards AJ. 1997. Plant breeding systems, 2nd edn. London: Chapman & Hall. Robertson C. 1928. Flowers and insects. XXV. Ecology 9: 505±526. Sasikumar B, George JK, Ravindran PN. 1992. Breeding behaviour of black pepper. Indian Journal of Genetics 52: 17±21. Semple KS. 1974. Pollination in Piperaceae. Annals of the Missouri Botanical Garden 61: 868±871. Smithe FB. 1975. Naturalist's color guide. New York: The American Museum of Natural History. Stelleman P, Meeuse ADJ. 1976. Anthecological relations between reputedly anemophilous ¯owers and syrphid ¯ies I. The possible role of syrphid ¯ies as pollinators of Plantago. Tijdschrift voor Entomologie 119: 15±31. Tebbs MC. 1989a. The climbing species of New World Piper (Piperaceae). Willdenowia 19: 175±189. Tebbs MC. 1989b. Revision of Piper (Piperaceae) in the New World. 1. Review of characters and taxonomy of Piper section Macrostachys. Bulletin of the British Museum of Natural History (Botany) 19: 117±158. Tebbs MC. 1993. Piperaceae. In: Kubitzki K, Rohwer JG, Bittrich V, eds. The families and genera of vascular plants. Berlin: SpringerVerlag. Waldbauer GP. 1984. Mating behavior at blossoms and the ¯ower association of mimetic Temnostoma spp. (Diptera, Syrphidae) in northern Michigan. Proceedings of the Entomological Society of Washington 86: 295±304. Weberling F. 1992. Morphology of ¯owers and in¯orescences. Cambridge: Cambridge University Press. Wright SJ. 1991. Seasonal drought and the phenology of understorey shrubs in a tropical moist forest. Ecology 72: 1643±1657. Yuncker TG. 1972. The Piperaceae of Brazil I: PiperÐGroup I, II, III, IV. Hoehnea 2: 19±366. Yuncker TG. 1973. The Piperaceae of Brazil II: PiperÐGroup V; Ottonia; Pothomorphe; Sarcorhachis. Hoehnea 3: 29±284. Yuncker TG. 1975. The Piperaceae of Brazil IV: Index. Hoehnea 5: 123±145. Zar JH. 1996. Biostatistical analysis, 3rd edn. New Jersey: Prentice Hall. Zeisler M. 1938. UÈber die Abgrenzung der eigentlichen Narben¯aÈche mit Hilfe von Reaktionen. Beihefte zum Botanischen Central Blatt 58: 308±318.
APPENDIX The following species were recorded and included in this study (herbaria numbers in parentheses): Piper aduncum Linnaeus (UEC 35299, IAC 31154) P. amalago (Jacq.) Yuncker (UEC 35300, IAC 31151) P. arboreum Aublet (UEC 35301, IAC 31155) P. crassinervium H.B.K. (UEC 35302, IAC 31153) P. gaudichaudianum Kunth (UEC 53303, IAC 31150) P. glabratum Kunth (UEC 35304, IAC 31146) P. macedoi Yuncker (UEC 35305, IAC 31147) P. mikanianum (Kunth) Steudel (UEC 35306, IAC 31149) P. mollicomum (Kunth) (UEC 35307, IAC 31156) P. regnelli (Miq.) C. DC. (UEC 35308, IAC 31148) P. xylosteoides (Kunth) Steud (UEC 35309, IAC 31152) Ottonia martiana Miq. (UEC 35311, IAC 31161) O. propinqua (Kunth) (UEC 35310, IAC 31162) Pothomorphe umbellata (L.) Miq. (UEC 30614)
Pollination Biology of Piperaceae Species in Southeastern Brazil RO D O L FO A N T OÃ N I O D E F I G U E I R E D O *{ and MA R L I E S S A Z I M A{ {Departamento de CieÃncias, Faculdade de CieÃncias e Letras Padre Anchieta, Rua Bom Jesus de Pirapora 140, 13207-660 JundiaõÂ, SaÄo Paulo, Brazil and {Departamento de BotaÃnica, Universidade Estadual de Campinas, 13083-970 Campinas, SaÄo Paulo, Brazil Received: 19 August 1999 Returned for revision: 22 October 1999 Accepted: 2 December 1999 Phenology, ¯oral biology and breeding system are described for 14 Piperaceae species at two sites covered by semideciduous forest in southeastern Brazil. Flowering occurred continuously throughout the year, with a peak during the windy months. All species had minute odorous and nectarless ¯owers visited by insects; ®ve of them showed a substantial degree of self-compatibility, and one was andromonoecious. Seven species were wind pollinated, and three were exclusively pollinated by insects. Flower visitors were mainly hover¯ies and bees. # 2000 Annals of Botany Company Key words: Piper spp., Ottonia spp., Pothomorphe umbellata, pepper family, phenology, ¯oral biology, breeding system, ambophily, semi-deciduous forest.
I N T RO D U C T I O N The family Piperaceae is pantropical with species distributed in the New World from Mexico to southeastern Argentina. Brazilian forests harbour 283 species of Piper L., 25 species of Ottonia Sprengel, and two species of Pothomorphe Miquel (Yuncker, 1972, 1973, 1975). In spite of the importance of the pepper family in tropical forests, the biology of its species is almost unknown in Brazil. Until recently little was known about the pollination biology of the paleotropical Piper nigrum L. (black pepper), the most well known of the Piperaceae due to its economic importance (Pouvreau, 1984; Richards, 1997). Martin and Gregory (1962) recorded wind as the most important pollen vector in black pepper, while pollination of neotropical Piper species is thought to be carried out by bees, beetles and drosophilid ¯ies (Janzen, 1991; McDade et al., 1994; Kricher, 1997), mainly on account of the studies made in Central America (Semple, 1974; Fleming, 1985). In South America, there have been no studies on the pollination biology of Piperaceae species. The present study describes ¯oral morphology and the breeding and pollination systems of 11 species of Piper, two of Ottonia, and one of Pothomorphe in a semi-deciduous forest of southeastern Brazil. Pollination biology is discussed in the light of what is known for black pepper and the Central American Piper species. M AT E R I A L S A N D M E T H O D S Study area and plant species Fieldwork was undertaken from 1 Mar. 1994 to 31 Dec. 1995 at the Reserva de Santa Genebra (228490 S, 478070 W, * For correspondence. E-mail roro®[email protected]
0305-7364/00/040455+06 $35.00/00
670 m alt.) and at the EstacËaÄo Experimental de JundiaõÂ /IAC (238060 S, 468550 W, 715 m alt.), two reserves of semideciduous forest in southeastern Brazil ( for geographical and vegetation descriptions see Morellato, 1992; Morellato and LeitaÄo-Filho, 1995). There is a marked seasonality in the study area, with a dry, cold season from April to September and a wet, warm season from October to March (Mello et al., 1994). The wind velocity increases at the end of the dry season (September). Piperaceae species were surveyed by transects made in the dry and swampy parts of the reserves. Species names follow Yuncker (1972, 1973), and voucher specimens were deposited at the herbaria of Universidade Estadual de Campinas (UEC) and Instituto AgronoÃmico de Campinas (IAC) (see Appendix). Phenology The phenology was registered through weekly observations on ®ve individuals of each Piperaceae species, from 1 Jan. to 31 Dec. 1995. The relationship between ¯owering periods and climate variables was tested using Spearman, correlation coecient, rs (Zar, 1996). Daily observations were carried out on in¯orescences to verify the period of anthesis and the temporal separation of stigma receptivity and anther dehiscence (types of dichogamy following Lloyd and Webb, 1986). Flower characteristics Flower morphology was described using ten in¯orescences of each species. Names of ¯oral parts follow Radford et al. (1974) and Weberling (1992). Colour names follow Smithe (1975). Stigma receptivity was checked with the hydrogen peroxidase reaction (Zeisler, 1938). # 2000 Annals of Botany Company
De Figueiredo and SazimaÐPollination in Piperaceae
Osmophores and nectaries were tested with neutral red 1 : 1000 for odour (Dafni, 1992) and 1 : 10 000 for nectar (Kearns and Inouye, 1993). Breeding system In order to analyse the occurrence of within-in¯orescence fertilization (autogamy and geitonogamy), 30 immature in¯orescences were isolated in waterproof bags, excluding pollen from other plants. Thirty unbagged, randomly chosen in¯orescences of the same age were left untreated as controls. The number of fruits that set and survived until day 20 after the ¯owering period was checked and used as an index of successful pollination. Pollination system Wind pollination experiments were conducted by isolating 30 immature in¯orescences in thin cloth bags that completely excluded visitors. Thirty unbagged in¯orescences of the same age were observed as controls. Fruit set was checked 20 d after ¯owering and used as an index of successful pollination. Open ¯owers of each Piperaceae species were observed for 60 h, from 0600 to 1800 h for veri®cation of insect visitors. The insects collected were identi®ed and deposited at the Identi®cation Centre of Phytophagous Insects (CIIF) of Universidade Federal do ParanaÂ. The behaviour of insect species was monitored by ®eld observations and the presence of pollen on their body parts was checked with a stereoscopic microscope. R E S U LT S A N D D I S C U S S I O N The plants and their phenology
Flower characteristics The Piperaceae in¯orescences show a variable number of minute ¯owers (Table 1, Fig. 2A), which are creamy, yellowish or whitish in colour (Table 1). These colours are also the most common for the Piperaceae in Central America (Tebbs 1989a, b), but Tebbs found some Piper species with pink and violet in¯orescences. Almost all species produce a sweet/lemon-lime odour, except for P. aduncum whose ¯owers are scentless. Lemon-lime odour occurs in some Mesoamerican Piperaceae species (Tebbs, 1989a). This odour is probably the most widespread in the family. Nectar is not discernible in the ¯owers, as has been observed for other Piper species (Fleming, 1985; Marquis, 1988; Ollerton, 1996). The ¯owers are astylocarpellous in Piper gaudichaudianum, P. glabratum, P. macedoi and Pothomorphe umbellata, astylocarpepodic in Ottonia species, and stylocarpellous in the other Piperaceae species. The style is absent in Piper gaudichaudianum, P. glabratum and P. macedoi, as well as in the Ottonia and Pothomorphe species. All other species are homostylous. The stigmas are decurrent in almost all species, except for P. amalago and P. arboreum, whose stigmas are clavate. Piper and Pothomorphe species commonly show three stigmas, while Ottonia species show four stigmas. The stamens are ®lantherous, their number varying from three to six in the species studied (Table 1). The stamens of Piper arboreum number four per ¯ower, but Tebbs (1989a) found six stamens in Central American populations. All Piperaceae species have divergent stamen thecae, with extrorse longitudinal dehiscence of the anthers, which drop after the pollination period. Pollen of all species is dry and of the monad type, white or yellow coloured.
12
45
10
40 35
8
30 25
6
20
4
15
Wind velocity
Most of the Piperaceae studied are shrubs, with the exception of Piper arboreum, which is a treelet, and P. mikanianum, which has an herbaceous habit. Piper gaudichaudianum and Ottonia martiana are found only in swamps, while the other species grow both in and at the edge of swamps and in dry parts of the semi-deciduous forest. The phenological pattern of the studied Piperaceae is of the continual type (sensu Newstron et al., 1994). The peak of ¯owering species coincides with the peak of wind velocity in the area (Fig. 1), although a strong correlation was not veri®ed (rs 0.56, 0.05 5 P 5 0.10). Some climatic variables have been correlated with the ¯owering periods of Central American Piper species (Opler et al., 1980; Marquis, 1988; Wright, 1991), but such relations were not observed in the species studied here. The proximal ¯owers in the in¯orescences open ®rst in the majority of species, with the exception of Piper mikanianum, P. xylosteoides and Pothomorphe umbellata, whose ¯owers all open at the same time. Stigma receptivity, as well as anther dehiscence, last up to 6 d in all species. In¯orescences are in bloom for up to 10 d, while Marquis (1988) found a ¯owering time of 1 week for the Mesoamerican species Piper arieianum C.DC.
The majority of the studied species exhibits incomplete protogyny, but Piper mikanianum shows complete protogyny, P. xylosteoides incomplete protandry, and P. regnelli homogamy. The primary type of dichogamy found in Piperaceae is incomplete protogyny, which has also been recorded for paleotropical species (Menon, 1949; Martin and Gregory, 1962).
No. of species flowering
456
10
2
5
0
0 J
F M A M J
J
A
S
O N D
Months FIG. 1. Number of Piperaceae species ¯owering (bars) and wind velocity (line) in semi-deciduous forest of southeastern Brazil in 1995.
De Figueiredo and SazimaÐPollination in Piperaceae
457
T A B L E 1. In¯orescence and ¯ower characteristics of Piperaceae species in semi-deciduous forest of southeastern Brazil Species Piper aduncum P. amalago P. arboreum P. crassinervium P. gaudichaudianum P. glabratum P. macedoi P. mikanianum P. mollicomum P. regnelli P. xylosteoides Ottonia martiana O. propinqua Pothomorphe umbellata
Number of ¯owers (x + s.d.) 505.00 + 19.13 351.40 + 28.50 403.60 + 18.23 544.00 + 18.51 407.20 + 14.54 446.00 + 53.20 607.20 + 19.54 199.20 + 8.29 577.20 + 54.21 746.20 + 40.06 290.20 + 33.12 118.80 + 2.59 284.60 + 18.05 449.00 + 39.40
Colour
Odour intensity
Length of the style (mm)
Length of the ®lament (mm)
Stamen number
±
0.5 1.5 1.0 2.0 sessile sessile sessile 0.5 0.5 1.0 0.2 sessile sessile sessile
0.5 3.0 4.0 2.4 0.3 0.5 0.5 1.0 1.0 1.2 0.2 0.8 1.0 ±
6 4 4 4 3 4 4 3 3 3 3 4 4 2
Cream Pale horn Sulphur yellow Spectrum yellow Sulphur yellow Pale horn Sulphur yellow Sulphur yellow Pale horn Straw yellow Cream diluted Sulphur yellow Sulphur yellow Pale horn
Some of these androecium features have previously been described for Piper (Yuncker, 1972, 1973; Endress, 1994). Breeding systems Most of the Piperaceae species are hermaphrodite, with the exception of Piper arboreum which has some in¯orescences with only staminate ¯owers and others with hermaphrodite ones. Andromonoecy has not previously been recorded in Piperaceae (see Tebbs, 1993, for a review of the family characteristics), although Ollerton (1996) noticed an individual of a monoecious Australian Piper species bearing only male in¯orescences. The ratio of staminate to hermaphrodite in¯orescences is 0.53 in P. arboreum, which shows the lowest fruit formation amongst the Piperaceae (Table 2). Five species show a substantial degree of selfcompatibility: Piper aduncum, P. mollicomum, P. regnelli, P. gaudichaudianum and Pothomorphe umbellata. Piper macedoi and P. xylosteoides set few fruits, suggesting that they manifest an unimportant degree of self-compatibility or that the bag initially contained few very small insects (such as thrips) carrying pollen from another plant. The other Piperaceae species are necessarily outcrossed (Table 2). As the microclimate inside the waterproof bags may have been very dierent to that outside (see Corbet and Willmer, 1981), and minute insects such as thrips may have been present inside them, the estimation of self-pollinated fruit development (Table 2) is conservative. The breeding systems of Piperaceae are known for only very few species: the paleotropical Piper nigrum and the neotropical P. arieianum which are self-compatible (Martin and Gregory, 1962; Marquis, 1988; Sasikumar et al., 1992), and the paleotropical P. methysticum which is selfincompatible (Prakash et al., 1994). Although tests for agamospermy were not carried out, Piper aduncum and Pothomorphe umbellata may be agamospermous, since all in¯orescences of these species developed fruits. It has been suggested that agamospermy occurs in Piper nigrum (Gentry, 1955).
Pollination systems Wind pollination occurs in seven of the studied Piperaceae: Piper arboreum, P. gaudichaudianum, P. macedoi, P. mikanianum, P. xylosteoides, Ottonia martiana and O. propinqua (Table 2). Anemophily has previously been recorded only for P. nigrum in Panama (Martin and Gregory, 1962), whereas other Central American Piper species are not wind pollinated (Semple, 1974; Fleming, 1985). As the bags used for insect exclusion may have aected access by windblown pollen, the estimation of wind pollination (Table 2) may be conservative. Furthermore, small insects initially present in the bag may have also contributed to fruit set by autogamy or geitonogamy, as perhaps occurred in P. crassinervium. Four species, P. aduncum, P. mollicomum, P. regnelli and Pothomorphe umbellata, did not yield substantially more fruits in insectproof bags than could have been accounted for by withinin¯orescence fertilization in waterproof bags, hence they were not considered to be wind pollinated. The majority of species grow at the edge of the forest, an appropriate place for pollen transport by wind (Arruda and Sazima, 1988), but Piper arboreum, P. gaudichaudianum, P. mikanianum, P. regnelli, O. martiana and O. propinqua also grow in the forest under-storey. These individuals, too, are eectively pollinated by wind (Table 2). Bawa and Crisp (1980) and Bawa et al. (1985) also recorded some anemophilous under-storey species within the Moraceae. Species of Piperaceae exhibit some classic traits of anemophilous plants, such as the increase of individuals ¯owering in windy months ( from September to December, see Fig. 1), the small and pulverulent pollen grains and the protogynous ¯owers (Lloyd and Webb, 1986; Honig et al., 1992). Piper amalago, P. crassinervium and P. glabratum may be exclusively insect-pollinated. As insects visit in¯orescences of all Piperaceae species (Table 2), these plants may well bene®t from insect pollination, although we have no evidence of this since none of our treatments excluded wind pollination. Flies (mainly hover¯ies, Syrphidae: Diptera) and bees (mainly Apidae: Hymenoptera) were the most frequent visitors, transporting Piperaceae pollen
458
De Figueiredo and SazimaÐPollination in Piperaceae
FIG. 2. A, In¯orescence of Ottonia martiana showing its minute ¯owers. A small hover¯y, Ocyptamus sp. (Diptera: Syrphidae), length approx. 8 mm, is visiting the in¯orescence. B, Copestylum tripunctatum (Syrphidae), 15 mm in length, visiting a hermaphrodite in¯orescence of Piper arboreum.
on their bodies (Fig. 2B). Central American Piper species are pollinated mainly by stingless bees of the genus Trigona (Apidae: Hymenoptera) (Semple, 1974; Fleming, 1985). Hover¯ies are common and generalist ¯ower visitors, being attracted by creamy-coloured and scented ¯owers,
even of anemophilous plant species (Robertson, 1928; van der Goot and Grabrandt, 1970; Stelleman and Meeuse, 1976; Waldbauer, 1984; Barth, 1991; Arruda and Sazima, 1996). Hover¯ies and bees visit Piperaceae ¯owers for pollen. Moving along the in¯orescence, the ¯ies touch the
De Figueiredo and SazimaÐPollination in Piperaceae
459
T A B L E 2. Number of Piperaceae in¯orescences pollinated by sel®ng, wind and open pollination (control), as well as receiving insect visitors, in semi-deciduous forest of southeastern Brazil Within-in¯orescence fertilization
Open pollination (control)
Wind pollination
Open pollination (control)
30
28
30
30
P. amalago
0
22
0
24
P. arboreum
0
6
8
7
P. crassinervium
0
15
1
13
P. gaudichaudianum
6
20
23
19
P. glabratum
0
15
0
18
P. macedoi
2
18
15
15
P. mikanianum
0
14
12
12
P. mollicomum
17
22
14
23
P. regnelli
30
27
23
28
P. xylosteoides
2
6
12
8
Ottonia martiana
0
24
27
25
O. propinqua
0
30
30
30
16
16
12
12
Species Piper aduncum
Pothomorphe umbellata
open anthers with the labell and gather pollen with the pulvilli and arolia. In some instances, hover¯ies also touch the bracts with the labell, presumably collecting pollen grains or some other substance. The bees move faster along the in¯orescence, collecting pollen with their legs. Flies visit several in¯orescences before leaving the plant, whereas bees visit only two to four in¯orescences, both of them moving from the proximal part of the in¯orescence to the distal part. The behaviour of these visitors may increase autogamy and geitonogamy (Barth, 1991; Richards, 1997). Species of Coleoptera and Hemiptera were also found in Piperaceae in¯orescences, but they rarely moved between ¯owers and behaved as herbivores, chewing and sucking ¯owers and fruits. Thomisidae spiders prey upon bees and ¯ies visiting the in¯orescences of Piper crassinervium and P. regnelli.
CO N C L U S I O N S This study has identi®ed ambophily (wind and small insects) as the primary pollination system for southeast Brazilian Piperaceae species. This is a well-based hypothesis, since all Piperaceae species show attributes commonly associated with anemophily and entomophily, but one which had not
Visiting insects
Number of visits (%)
Diptera Hymenoptera Lepidoptera Diptera Hymenoptera Diptera Hymenoptera Diptera Hymenoptera Lepidoptera Diptera Hymenoptera Diptera Hymenoptera Lepidoptera Diptera Hymenoptera Diptera Hymenoptera Diptera Hymenoptera Diptera Hymenoptera Lepidoptera Diptera Hymenoptera Diptera Hymenoptera Diptera Hymenoptera Lepidoptera Diptera
255 8 1 241 56 165 36 663 41 5 16 5 131 15 1 75 10 95 17 103 8 1500 604 2 47 9 35 2 81 106 1 8
(96.59) (3.03) (0.38) (81.21) (18.79) (82.09) (17.92) (92.86) (5.78) (0.71) (76.19) (23.81) (89.12) (10.20) (0.68) (88.24) (11.76) (84.82) (15.18) (92.79) (7.21) (71.23) (28.68) (0.09) (87.04) (16.07) (94.74) (5.26) (43.09) (56.38) (0.53) (100.00)
previously been demonstrated experimentally. Besides the continual ¯owering phenology, the incomplete protogyny, the odorous in¯orescences and the self-pollination, the study detected an andromonoecious species, the ®rst record of andromonoecy for this plant family.
AC K N OW L E D G E M E N T S The authors thank Conselho Nacional de Desenvolvimento Cientõ ®co e TecnoloÂgico (CNPq, process number 140055/ 94-0) for ®nancial support. This work was presented at the XVI International Botanical Congress (Saint Louis, USA) and R. A. de Figueiredo also acknowledges fellowships from FundacËaÄo de Amparo aÁ Pesquisa do Estado de SaÄo Paulo (FAPESP, process number 1999/04469-8) and from XVI IBC Fellowship Committee. Insect identi®cations were carried out by L. Marinoni, Pe. J. S. Moure, C. Carvalho, L. M. Almeida, O. Mielke and K. Zanol. FundacËaÄo Jose Pedro de Oliveira and EstacËaÄo Experimental de Jundiaõ / IAC provided permissions to work on their forest reserves. We are indebted to S. Corbet and D. Inouye for providing valuable comments and suggestions that improved an earlier version of the paper.
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APPENDIX The following species were recorded and included in this study (herbaria numbers in parentheses): Piper aduncum Linnaeus (UEC 35299, IAC 31154) P. amalago (Jacq.) Yuncker (UEC 35300, IAC 31151) P. arboreum Aublet (UEC 35301, IAC 31155) P. crassinervium H.B.K. (UEC 35302, IAC 31153) P. gaudichaudianum Kunth (UEC 53303, IAC 31150) P. glabratum Kunth (UEC 35304, IAC 31146) P. macedoi Yuncker (UEC 35305, IAC 31147) P. mikanianum (Kunth) Steudel (UEC 35306, IAC 31149) P. mollicomum (Kunth) (UEC 35307, IAC 31156) P. regnelli (Miq.) C. DC. (UEC 35308, IAC 31148) P. xylosteoides (Kunth) Steud (UEC 35309, IAC 31152) Ottonia martiana Miq. (UEC 35311, IAC 31161) O. propinqua (Kunth) (UEC 35310, IAC 31162) Pothomorphe umbellata (L.) Miq. (UEC 30614)