Cipadessa baccifera populations growing at Sunkarametta (18.2783° N, 82.9670° E) and Kasipatnam (18.1732° N, 83.0771° E) near Ananthagiri Reserve Forest in Alluri Sitharama Raju District, Andhra Pradesh, India, were used for the present study during June 2021 to December 2022. Field observations were made to record flowering period, floral biology, foraging activity and pollination, breeding systems, and fruiting and seed dispersal aspects. Fifty flowers from 20 plants were collected to describe morphological aspects of flowers briefly because of reports of inconsistencies on these aspects in the literature. Fifty mature buds tagged on 10 plants were followed for recording the timing of anthesis and anther dehiscence. A 10X hand lens was used to confirm the dehiscence time and mode. Stigma receptivity was observed by using H₂O₂ test described in Dafni et al. (2005).

Twenty-five mature buds which were about to open were bagged before sunrise and removed at noon time to measure total nectar produced by each flower by inserting a micropipette into the flower base. Since nectar was not secreted after noon time, the flowers were not monitored thereafter during the entire period of their lifespan. The average volume of nectar produced by all these flowers was taken as the total volume of nectar/flower and expressed in µl. The nectar produced in these flowers was used for measuring nectar sugar concentration and then for calculating the mean sugar concentration. A hand sugar refractometer (Erma, Japan) was used for measuring nectar sugar concentration. Nectar analysis was carried out for sugar types and amino acid types using the paper chromatography method described in Dafni et al. (2005).

Twenty mature but undehisced anthers were collected from different plants and placed in a Petri dish. Later, each time a single anther was taken out and placed on a clean microscope slide and dabbed with a needle in a drop of lactophenol-aniline blue. The anther tissue was then observed under the microscope for pollen. The pollen mass was drawn into a band and the total number of pollen grains was counted under a compound microscope (400X). This procedure was followed for counting the number of pollen grains in each anther collected. Based on these counts, the mean number of pollen grains produced per anther was determined. The mean pollen output per anther was multiplied by the number of anthers in the flower for obtaining the mean number of pollen grains per flower. Another set of 10 dehisced anthers was collected in a Petri dish and the pollen removed from these anthers was examined under microscope for recording the pollen grain features.

The protocols described by Mondal et al. (2009) were followed for identifying amino acid types present in the pollen. Pollen was collected from mature anthers and filtered through sieving using meshes of different size (100, 200 and 300 µm) to remove the debris. Then, the pollen was rapidly dried over silica gel at 30°C and stored. Free amino acids were extracted from the pollen using the method of Bieleski and Turner described in Mondal et al. (2009). Later, the extract thus obtained was used for the qualitative analysis of the free amino acids of pollen using thin layer chromatography.

Field observations were made on the foraging activity of insects before sunrise, after sunset and during daylight hours. The foraging schedule, forage collected and the flower probing behavior of each insect species was recorded. The number of foraging visits made by each insect species was recorded for 15 min at each hour during the entire length of the observation period to examine the pattern of foraging activity according to the availability levels of nectar and pollen. This field observation on foraging activity of insect species was repeated on four clear sunny days and the data thus collected were used to calculate the average number of visits made by each species at each hour of the day and the percentage of foraging visits by each category of insect species to record the foraging rate of individual insect species and each insect category. The foraging behavior of each insect species was observed with reference to its approach, landing, probing behavior employed for pollen collection and contact with essential organs in effecting pollination.

Breeding systems were tested for different modes of self-pollination, and cross-pollination. Spontaneous autogamy was tested by bagging twenty flowers without hand-pollination. Twenty mature buds were bagged, opened the next day after the occurrence of anthesis, anther dehiscence and stigma receptivity; the stigma was pollinated with the pollen of the same flower using a brush and bagged again to test hand self-pollination. Twenty mature buds were bagged after emasculation, opened the next day after the commencement of stigma receptivity; the stigma was pollinated with the fresh pollen of a different flower of the same plant using a brush and again bagged to test geitonogamy. Twenty mature buds were bagged after emasculation, opened the next day after the occurrence of stigma receptivity; the stigma was pollinated with the fresh pollen from the flowers of a different plant using a brush and again bagged to test xenogamy. The bagged flowers were followed for 30 days for fruit set. Based on the flowers that produced fruits, the percentage of fruit set was calculated. Five flowers each from 10 plants were tagged prior to anthesis and followed for fruit set in open-pollinations. The percentage of fruit set was calculated based on the number of fruited flowers. Fruits are 5–10 seeded but seed set rate was not calculated in any mode of pollination.

Fruit and seed characters and seed dispersal

Fruit and seed characters were described in view of the reports of inconsistencies on these characters. Field observations were made on fruit dispersal agents and the role of dispersal agents was briefly described. Brief field observations were made on the production of new plants from seed and perennial old root stock to record vegetative growth and development.

Cipadessa baccifera is a seasonal woody shrub which grows on hill slopes and hill sides with rocky bed (Fig. 1A). With the onset of wet season in June, the underground stout root stock sprouts to produce new growth and begin sexual reproduction first (Fig. 1B). Seeds from the soil bank also germinate at the same time to produce new plants and add new individuals. These new individuals show sexual reproduction about two months later to the plants that emerged from the old root stock. The dual mode of propagation functional in this plant contributes to the extension of the period of sexual reproduction. The flowering occurs early in plants that originated from the previous root stock and late in plants that emerged from seeds. The flowering season begins from July and ends in November but it extends into January provided the soil is sufficiently wet. The aerial parts gradually wither away by the end of March. The inflorescence is an axillary cymose inflorescence consisting of few-clustered flowers (Fig. 1C).

The flowers are pedicellate, creamy-white, 2 mm long, 3–4 mm diameter, actinomorphic and bisexual (Fig. 1E). The calyx is 5-lobed, each lobe 2 mm long, pubescent, triangular and spreading. The corolla has five free creamy white petals, and each petal is ovate and 3–4 mm long. The stamens are 10 with filaments connate to the extent of ¾ of their length forming an urceolate staminal tube with hairy middle portion (Fig. 1F, G); the filaments are terminated by a pair of lanceolate erose appendages. The anthers are yellow, acute, pubescent and inserted just below the appendages. The ovary has patelliform disk with 5–6 locules and each locule has two collateral ovules. The style is short, stout and columnar with swollen capitate to turbinate stigma consisting of 5–6 stigmatic lobes without any hairs.

Mature buds are open during 07:30–09:30 h (Fig. 1C). The process of bud opening involves only the unfolding of petals exposing the staminal tube with anthers inside, which lie on the rim of capitate portion of the style-head throughout flower life (Fig. 1D, E). The anthers dehisce by longitudinal slits shortly before the petals unfold and the pollen from anthers is attached to the rim of capitate style-head which is unreceptive to pollen during the entire period of flower life (Fig. 1F, G). The turbinate portion of the style-head protrudes out from the style at 15:00 h, is receptive to pollen almost instantaneously and its receptivity continues until the evening of the following day. The pollen grains are 375 ± 36 per anther and 3,750 per flower. The pollen grains are prolate-spheroidal, 35–40 µm, tetra-colporate with smooth exine and thickened apertures. The pollen contains four essential amino acids which include threonine, methionine, lysine and arginine, and five non-essential amino acids such as alanine, amino-butyric acid, cystine, glycine and hydroxy proline (Table 1). Nectar is secreted in minute volume at the base of ovary and concealed by the staminal tube; its secretion ceased about 2 h after anthesis. It is accessible to nectar foragers only when they probe through the narrow gaps between staminal sheath and the columnar style. Individual flowers produce 1.2 ± 0.45 µl of nectar with sucrose, glucose and fructose sugars; the sugar concentration was 25 ± 2.5%. The nectar contains six essential amino acids such as threonine, valine, leucine, isoleucine, histidine and arginine, and nine non-essential amino acids such as alanine, amino-butyric acid, cysteine, cystine, glutamic acid, glycine, hydroxy proline, proline and serine (Table 1). The petals together with the staminal tube wither away and fall off on 4th day while the columnar style and its head falls off on 9th or 10th day, and the sepals gradually dry up. The ovary portion gradually grows and bulges out to produce fruit.

In open flowers, the style-head is capitate without any projection from its centre and this portion is non-receptive to pollen. But, turbinate head with stigmatic lobes springs out from the centre of capitate style-head from 15:00 h onwards on the day of anthesis. The stigmatic lobes commence receptivity to pollen almost spontaneously and continue to be receptive until the evening of the next day. The anthers lie on the capitate style-head throughout the flower lifespan. Since the anthers dehisce during bud stage, the pollen grains attach automatically to the the rim of capitate style-head which is non-receptive to pollen and hence, it merely acts as pollen presenter and facilitates only the occurrence of cross-pollination until the time of emergence of turbinate style head with its stigmatic lobes by which time most of the self-pollen is picked up by the pollen vectors. However, the temporal separation of male and female phases displayed by protandry and secondary pollen presentation do not totally prevent the occurrence of vector-mediated autogamy and geitonogamy.

The flowers were visited by insects during day time from 09:00 to 16:00 h with a peak of foraging activity during 10:00–12:00 h (Fig. 2). The insects visiting the flowers were one honey bee species, Apis cerana F. (Apidae) (Fig. 3A), one halictid bee species, Nomia sp. (Halictidae) (Fig. 3B), four wasp species, Polistes sp. (Fig. 3C), Rhynchium sp. (Fig. 3D) (Vespidae), Sphex sp. (Fig. 3E), Eremnophila sp. (Fig. 3F) (Sphecidae), and three lycaenid butterflies, Chilades pandava Horsfield (Fig. 3G), Spindasis vulcanus F. (Fig. 3H) and Zizeeria karsandra Moore (Fig. 3I) (Table 2). All these insects collected nectar while the bees also collected pollen. They approached the flowers in upright position, landed on the petals and then probed the flowers for nectar by inserting the tongue/proboscis through the narrow gap between the appendages of the staminal tube and the style-head. Since the staminal tube is short, these insects accessed the nectar location with great ease. All these insects collected pollen and/or nectar during unreceptive and receptive stigma phases of flowers; pollen was carried away without pollination effect by them during unreceptive phase of the style-head while pollen transfer and pollination occurred during receptive phase of the style-head. These insects approached the flowers in upright manner, landed on the urceolate staminal tube and inserted their tongue/proboscis through the narrow gaps between the staminal tube and style-head to collect the nectar seated at the ovary base. In flowers with the unreceptive phase of the style-head, the forehead and tongue/proboscis/ventral side of abdomen of insects invariably contacted the rim of style-head lined with pollen. In consequence, the pollen was transferred to these parts of the insects and the pollen thus carried away by the insects is transferred to other conspecific flowers in their successive foraging visits for nectar and/or pollen collection. Pollination occurs only when they visit the flowers with receptive style-head. In flowers with receptive style-head, these insects first contacted the turbinate portion of the style-head with their forehead and the ventral side of their body; then pollen transfer to the receptive portion of the turbinate style-head occurred if the insect had visited the flowers previously and carried pollen on their dorsal side. As a result, either self- or cross-pollination occurs depending on the pollen source. The honey bee, Apis cerana, loaded the collected pollen into its pollen baskets. The alkali bee (Nomia sp.) is known to nest in salt-saturated soils. In flowers with unreceptive style-head, the bees gathered pollen from the rim of the capitate style-head and in this act, the bees moistened the pollen with the saliva using its tongue, transferred it to its hind legs and packed it into the pollen basket. The pollen basket is a particular arrangement of hairs that occur on the tibia of the hind legs of the bees. When the bees with pollen load made successive visits to the flowers with receptive style-head, its pollen basket brushed against the turbinate portion of the style-head which ended up in pollen transfer and pollination.

The temporal separation of male and female phases within the flowers does not facilitate the occurrence of both self- and cross-pollination until the time of commencement of receptivity of the turbinate style-head but provides opportunity for both self- and cross-pollination after turbinate style-head commences receptivity to pollen in the evening hours of day 1 flowers and day-long in day 2 flowers. But, insects did not visit the flowers during the receptivity period of style-head in day 1 flowers indicating that the day 1 flowers act as pollen donors while day 2 flowers principally act as pollen recipients. Such a strikingly protandrous condition appeared to prevent the occurrence of autogamy and allow geitonogamy and xenogamy. Among the insects recorded at the flowers, wasps and lycaenid butterflies were found to be visiting the flowers across conspecific plants within the population and also between nearby populations in quest of more nectar collection and such inter-plant foraging activities by them appeared to be promoting cross-pollination and reducing geitonogamous pollination. The bees being pollen and nectar foragers tended to spend more time at individual flowers and on the same plant, and such a foraging behavior was treated to be promoting geitonogamous pollination rather than cross-pollination. Of the total foraging visits made by all insect species, bees and wasps groups made 39% each and butterflies 22% (Fig. 4). Body washings of insects which were collected at 11:00 h for assessing pollen carry-over efficiency showed that the mean number of pollen grains carried by individual species ranged from 13.3 to 97.3. The bees carried a mean number of 83.8 to 97.3 pollen grains, wasps from 26.8 to 45.8 and butterflies from 13.3 to 34.3 (Table 3). These values indicated that the bee species had high pollen carrying capacity when compared to wasps and butterflies but bees are important for effecting mostly geitonogamous pollination. Wasps had moderate pollen carrying capacity but their frequent inter-plant foraging activity is important for effecting mostly xenogamous pollination. Butterflies are also important for promoting xenogamous pollination but they had low pollen carrying capacity. Therefore, wasps were found to be the principal pollinators in promoting cross-pollination while the bee and butterflies are supplementary pollinators.

Hand-pollination tests for breeding systems showed that spontaneous autogamy is not functional while manipulated autogamy, geitonogamy and xenogamy are functional but fruit set in manipulated autogamy is negligible as it stood at 5% only. Fruit set is 40% in geitonogamy, 80% in xenogamy and 82% in open-pollination mode (Table 4). The results of hand-pollination tests showed that the plant has facultative xenogamous breeding system which is completely vector-dependent.

Fruits mature within 6–7 weeks and display color shifts from green to light red to dark and finally to reddish brown (Fig. 5A–F). Fruit is a fleshy globular drupe with five pyrenes each with one or two seeds, 5–8 mm across and all seeds are united by a common translucent fleshy arillode. The seeds are orange-segment-shaped with thin and membranous testa (Fig. 5G, H). The fruits are edible and eaten occasionally by 20 species of frugivorous birds belonging to 10 families of the order Passeriformes during day time from 08:00 to 16:00 h. They were Artamus fuscus Vieillot (Artamidae), Pericrocotus cinnamomeus Linnaeus, P. flammeus J. R. Forster (Campephagidae), Dendrocitta vagabunda Latham (Corvidae), Dicaeum erythrorhynchos Latham (Dicaeidae), Dicrurus adsimilis Bechstein, D. caerulescens Linnaeus (Dicruridae), Turdoides caudatus Dumont (Leiothrichidae), Oriolus kundoo Sykes, O. xanthornus Linnaeus (Oriolidae), Pycnonotus cafer Linnaeus, P. jocosus Linnaeus, P. luteolus Lesson (Pycnonotidae), Gracula religiosa Linnaeus, Sturnus roseus Linnaeus, S. malabaricus Gmelin, S. pagodarum Gmelin, Acridotheres ginginianus Latham, A. tristis Linnaeus (Sturnidae) and Tephrodornis pondicerianus Gmelin (Vangidae) (Table 5). These birds possess a short, curved or uncurved beak with a specialized tip for feeding on edible part of the fruits. They were found to open the fruits to feed on the fleshy arillode in which seeds are enclosed; the seeds were left out by many of these birds. They usually plucked the fruits, held them with their beak and carried them to new locations where they broke open the fruits to feed on the arillode and dropped the seeds. The seeds germinate during rainy season if the soil has sufficient moisture and nutrients. Since the birds carry them to new locations to feed, they are treated as effective seed dispersal agents. Further, when the birds fed on the arillode without plucking the fruits from the plant, the seeds fallen at the parental habitats germinate during rainy season adding new individuals to the existing population. The unutilized fruits fall to the ground and disperse by gravity and rain water easily because the habitats of the parental population are slopy and facilitate easy dispersal of fruits/seeds. The seeds become exposed after the decomposition of the composite transculent arillode and germinate during rainy season to produce new plants.