Journal of Insect Conservation 4: 203–210, 2000. © 2000 Kluwer Academic Publishers. Printed in the Netherlands.
Practical conservation paper
Importance of conserving alternative pollinators: assessing the pollination efficiency of the squash bee, Peponapis limitaris in Cucurbita moschata (Cucurbitaceae) Mar´ıa Azucena Canto-Aguilar∗ & V´ıctor Parra-Tabla Departamento de Ecolog´ıa, Facultad de Medicina Veterinaria y Zootecnia, Universidad Aut´onoma de Yucat´an, Km. 15.5 Carr. M´erida-Xmatkuil, Apartado postal 4-116, Itzimn´a C. P. 97000, M´erida Yucat´an, M´exico ∗ Author for correspondence (e-mail:
[email protected]; phone/fax: +52(99)423206, +52(99) 423206) Received 19 January 2000; accepted 20 April 2000
Key words: Apis mellifera, non-Apis bee conservation, pollination efficiency, Yucat´an, M´exico
Abstract Although the honey bee, Apis mellifera, has been considered the best pollinator for crops needing insect pollination, the current pandemic of varroatosis among honeybees highlights the need to find additional or alternative species as managed crop pollinators. Moreover, there is evidence that A. mellifera may not always be the most efficient pollinator. Introduction of A. mellifera into crops may be unnecessary, and even detrimental to non-Apis bee populations, which should be considered as an alternative for crop production improvement. Evaluating the pollination efficiency of non-Apis bees is one of the first steps in planning successful strategies for their conservation. In this study, we evaluated the pollination efficiency of Peponapis limitaris and A. mellifera in plots of Cucurbita moschata: pollen removal and deposition; pollinator visit frequency; and the pollinator visit–nectar production relationship. The results show P. limitaris to be the most efficient pollinator as: (1) both males and females remove and deposit almost four times as much pollen as A. mellifera; (2) they make significantly more floral visits than A. mellifera; and (3) their visit frequency shows a strong relationship to C. moschata nectar production during anthesis. Recommendations arising from this study are: (1) the introduction of A. mellifera be avoided in C. moschata crops; and (2) basic research be done on the biology of P. limitaris that contribute to its conservation and greater exploitation.
Introduction In addition to its use for honey and wax production in apiculture, the European bee, Apis mellifera, is widely used as a pollinator for the majority of cultivated species requiring insect pollination (Free 1970; McGregor 1976). Although the efficiency of Apis mellifera as a pollen vector for many cultivated species has not been evaluated (Stephen 1972), the introduction of hives into cultivated fields is a common agricultural practice. However, the current pandemic among honeybees caused by plagues such as Acaris woodi and Varroa jacobsoni (Oldroyd 1999),
JICO55 (BIO2FAM)
and africanisation, has not only increased the cost of Apis mellifera hive introduction and maintenance in agricultural zones, but also highlights the need to find alternative species as managed crop pollinators (Sugden 1993). Evidence also exists suggesting that native bees can be equal or better pollinators than Apis mellifera for certain agriculturally important species, such as Solanaceae (O’Toole 1993) and forage legumes (Richards 1996). Consequently, interest has grown in finding non-Apis pollinators that are potentially useful in agriculture, this being further stimulated by the importance of the ecological services of non-Apis pollinators to wild plant communities and agricultural crops
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(Thompson 1997; Allen-Wardell et al. 1998; Kearns et al. 1998). The search for non-Apis pollinators has exposed a gap in knowledge of the natural history of this type of pollinator. In order to establish successful management and conservation strategies for non-Apis pollinators, one of the first steps is to undertake detailed studies of their crop pollination efficiency. These should provide key information, such as the effect of this pollinator on fruit and seed production (Primack 1993; Torchio 1994). Bees of the genus Peponapis are an attractive option as alternative pollinators for squash crops such as Cucurbita pepo, C. moschata, C. ficifolia and C. mixta (Whitaker 1980; Tepedino 1981; Buchmann & Nabhan 1996). Given the close ecological relationship between the Peponapis and Cucurbita genera (Hurd et al. 1971), they may be the most adequate pollinators for these Cucurbita species. This is potentially significant because various important squash crops are used on the Yucatan Peninsula (Lira 1995), and are considered subsistence crops. Cucurbita species have formed a fundamental part of the diet and economy of the agricultural Mayan society in the region since the prehispanic era (Lira 1988; Nee 1990). Bees of the Peponapis genus are capable of obtaining nectar and pollen from other plant genera (Hurd and Linsley, 1964 in Hurd et al. 1971), however various important aspects of its life cycle depend on Cucurbita flowers (Hurd et al. 1971). Mating generally occurs in the flowers, and males take shelter in them during the day, once the flowers have begun to wilt. The aspect that most highlights the dependence of Peponapis on Cucurbita, and ensures its presence in squash crops, is that the female bees supply their larval cells with squash pollen and nectar, and are oligolectic on cucurbits (Hurd et al. 1971; Willis & Kevan 1995). However, this same dependence, coupled with non-aggressive behaviour towards other floral visitors, makes Peponapis vulnerable to competitors such as A. mellifera (Pinkus 1998). Because of the abundance of Cucurbita in cultivated areas it is common to observe A. mellifera and various Peponapis species foraging on its flowers (Mel´endez 1997). One comparative study has already been done of pollination efficiency in the Peponapis-Cucurbita system in the presence of A. mellifera (Tepedino 1981), though this study was done in areas where the Peponapis genus is considered introduced (Kevan et al. 1988). Research on this system is still needed in areas such as Yucatan, which are seen as centres of Cucurbita
domestication (Nee 1990; Hurd et al. 1971), and that have a high richness of Peponapis species (Mel´endez 1997). This study was done to evaluate specific aspects of A. mellifera and P. limitaris pollination efficiency in C. moschata crops on the Yucatan Peninsula. The proposed project objectives include determining (1) the most efficient pollinator in terms of inter-flower pollen removal and deposition; (2) if pollinator types differ in floral visit frequency; (3) if pollinators preferentially visit a certain floral type (pistillate and/or staminate); and (4) if there is a relationship, or synchronisation, between nectar availability patterns and pollinator foraging activity.
Materials and methods Study site The study was done between December 1995 and July 1996 during two planting periods: December 1995 and May 1996. The study site is located at Kilometre 15 of the M´erida-Xmatkuil highway (20◦ 580 North Latitude and 89◦ 370 West Longitude), and consists of three artificially irrigated, 12 × 32 m experimental plots. Climate in the zone is hot subhumid (Aw0) according to the Koppen classification as modified by Garc´ıa (1981), with an annual mean temperature of 26◦ C, and annual precipitation varying between 940 and 1132 mm (Duch 1988). The plots were surrounded by seasonally dry tropical forest secondary vegetation, including important nectar source species such Piscidia piscipula (Leguminoseae), and Viguiera dentata (Asteraceae). Using methods employed by agriculturists in the region, C. moschata seeds were sown in the plots. Eight seeds were placed in a 25 cm diameter by 25 cm deep hole, the soil being mixed with chicken faeces as a fertiliser. Each hole was located approximately 3 m from other holes, resulting in a total of 123 plants per plot. For each planting period, sampling was done during times when C. moschata was most actively flowering: mid-December 1995; all of February 1996; and from mid-May to early June 1996. Cucurbita moschata The squash species C. moschata is considered to be one of the oldest of its genus (Withaker 1980; Nee 1990), and has been cultivated on the Yucatan Peninsula
Importance of conserving alternative pollinators since the prehispanic era. It is presently one of the most important subsistence crops in the agricultural society of the region (Lira 1995; Zizumbo 1992). The plant is herbaceous, annual, and has creeping growth. Its growth cycle is short, lasting 1–3 months, and it flowers during approximately two and a half months. Generally, it grows in seasonal crops, although it can grow wild out of season, mixed with natural vegetation (Lira 1988). C. moschata is monoecious, which permits floral type estimation. The flowers themselves are a conspicuous yellow/orange colour, and of short duration (ca. 6 h), opening just before dawn. Both floral types have abundant nectar (see Results). Pistillate flowers can contain as many as 700 ovules, with an average of 393.75 ± 206.32 (x¯ ± DE, n = 16 fruits) produced per fruit (Ancona, unpublished).
Pollen removal Due to the morphological and behavioural differences between male and female P. limitaris (Hurd et al. 1971; Willis & Kevan 1995), all efficiency estimations were done for each sex. As a result, three ‘types’ of pollinators were evaluated: A. mellifera workers; P. limitaris males; and P. limitaris females. The quantity of pollen removed by each bee from C. moschata anthers was estimated by excluding a small group of staminate floral buds (n = 119 buds total) on a daily basis during peak flowering. When the floral buds opened, one pollinator was allowed to enter, which was subsequently captured and identified upon leaving the flower. This procedure was executed immediately after dawn, when individuals arriving at flowers are beginning foraging activities, thus diminishing the probability of overestimating the quantity of pollen removed. Each collected bee was preserved in a solution of 70% ethyl alcohol and liquid soap in order to facilitate the detachment of pollen stuck to its body. Once the pollen was obtained within the solution, a 9000 µl sample was taken. Ten subsamples of 60 µl each were then taken and the pollen grain content in each aliquot counted using a stereoscopic microscope (10×). Finally, the average number of pollen grains for the ten subsamples was multiplied by the total 9000 µl, and the result divided among the volume of each subsample (60 µl). This procedure was also applied to 25 unvisited flowers in order to estimate the pollen production of C. moschata.
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Pollen deposition Pollen deposition on the floral stigma was estimated by excluding 3 pistillate floral buds each day during peak flowering, just before anthesis, with a total of 95 buds excluded. When the flowers opened, a single bee visit was allowed, the bee being identified once inside the flower. After the bee visit, the flower was again covered and approximately 3 h allowed to pass for the deposited pollen to germinate (Stanley & Linskens 1974). The stigmas were then extracted and fixed in an alcohol– acetic acid solution, and stained with safranine, aniline blue, and acetic acid (Dafni 1992). Pollen grains with a pollinic tube (i.e. viable) were counted for each flower using a stereoscopic microscope (10×). Visit rate The number of visits per bee was estimated by counting legitimate visits, those with anther or stigma contact. Counts were made for 10 min periods with 5 min rests during the 6 h the C. moschata flowers were open, with a total of 24 h of observation over a 4 day period. A distinction was made between arrivals at pistillate and staminate flowers. Nectar production The nectar volume produced by each flower type was quantified during the anthesis (from 05:00 to 11:00 h). Over a 3 day period, exclusion bags were placed over 5 pistillate and 5 staminate flowers each day, for a total of 30 flowers. The nectar accumulated during 1 h periods was extracted with a graduated micropipette (10 and 15 µl) (Drummond Scientific), and the volume calculated using Cruden and Hermann’s (1983) method. Statistical analysis For pollen removal and deposition analysis the data were transformed with a square root to fulfil statistical normality requirements (Zar 1984), but the averages and respective standard errors (x¯ ± SE) are reported with untransformed values. An ANOVA was done to determine differences between pollinators, and a Tukey multiple comparative test was used when necessary to identify differences between pollinators. Pollinator visit frequencies were analysed using contingency tables, via a generalised lineal model (GLIM 4 1992) with Poisson-type random error distribution and linking logarithmic function (Crawley 1993). In this
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model the effect of pollinator type and C. moschata floral type as causal factors in floral visit frequency were evaluated. Nectar production data together with pollinator visit frequency data were transformed with a square root. Visit data between 1 h and another were combined in a single data point so as to make nectar production and visit frequency compatible, and to fit linear regressions for each pollinator type. When necessary, a comparison analysis was done between slopes (Zar 1984). Nectar production averages (x¯ ± SD) and floral visit frequencies are reported untransformed.
Results Pollen removal was different among pollinators (F = 19.5, p < 0.001), with P. limitaris females removing a significantly greater pollen quantity per visit (4879.7 ± 1308.3, n = 11); almost three times more than other pollinators (Figure 1A). This represents 19% of the average pollen production per C. moschata flower (25,385 ± 7698, n = 25), in a single visit. Male P. limitaris and A. mellifera workers removed statistically similar pollen quantities per visit (Figure 1A): an
average of 1608.2 ± 173.5 (n = 41) for male P. limitaris, representing 6% of pollen production; and 1282.9 ± 124.4 (n = 67) for A. mellifera workers, or 5% of pollen production. Pollen deposition on floral stigmas exhibited a pattern similar to pollen removal, with deposited pollen varying significantly among the three pollinators (F = 7.2, p = 0.001). Female P. limitaris transferred the greatest quantity of pollen on to stigmas (481.4 ± 98.2, n = 33), which is more than double that for A. mellifera, and three times that for male P. limitaris (Figure 1B). Average pollen deposition for the latter two pollinators was 253.4 ± 62.7(n = 42) for A. mellifera, and 177 ± 76.7(n = 20) male P. limitaris. Contingency table analysis showed that the quantity of visits to C. moschata flowers depended principally on pollinator type, although flower type had an evident impact on the bees (Table 1). The explained variation percentage of the model was 63%, with 89.52% of this corresponding to pollinator type and the remaining 10.48% to floral sex. P. limitaris males made more visits than the other pollinators and these visits concentrated on staminate flowers twice as frequently as on pistillate flowers. Generally, staminate flowers received more visits than the pistillate, except from female P. limitaris, which visited both floral sexes with equal frequency (Table 1). A. mellifera was the least frequent visitor to C. moschata, and its visits were principally to staminate flowers (Table 1). Pistillate C. moschata flowers produced an average of twice the nectar (119.70 ± 58.72, n = 16) than staminate flowers (47.99 ± 26.25, n = 19). Nectar production in the staminate and pistillate flowers begins moments after floral aperture, and though initially low, the quantity increases until reaching maximum production between 07:00 and 08:00 h. After this hour
Table 1. Chi-squared (χ 2 ) analysis of floral visits frequency for C. moschata. Expected frequencies are shown in parentheses. Flower sexa
Staminate Pistillate
P. limitaris males
P. limitaris females
A. mellifera workers
240 (220.35) 99 (118.65)
57 (75.40) 59 (40.60)
10 (9.75) 5 (5.25)
Significant differences between pollinators by flower sex; χ 2 = 17.36, p < 0.0001. a Significant differences between flower sexes; χ 2 = 44.84, p < 0.0001. b Significant differences between pollinators; χ 2 = 383.20, p < 0.0001. a,b
Figure 1. Averages (± SE) for (A) pollen removal from anthers and (B) pollen deposition on stigmas of C. moschata by A. mellifera (honey bees), and male and female P. limitaris (squash bees). The different letters indicate significant differences in averages (Tukey test p < 0.05).
Pollinatorb
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Table 2. Linear regression analysis between pollinator visit and nectar production through C. moschata anthesis. Pollinator
Lineal model
r2
F
p
n
P. limitaris males P. limitaris females A. mellifera workers
Y = 0.18 + 0.66X
0.31
11.69
0.0021
28
Y = −0.02 + 0.50X
0.65
48.73
