Published papers

The Chilean Puya species, Puya coerulea var. violacea and P. chilensis bear blue and pale-yellow flowers, respectively, while P. alpestris considered to be their hybrid-derived species has unique turquoise flowers. In this study, the chemical basis underlying the different coloration of the three Puya species was explored. We first isolated and identified three anthocyanins: delphinidin 3,3ʹ,5ʹ-tri-O-glucoside, delphinidin 3,3′-di-O-glucoside and delphinidin 3-O-glucoside; seven flavonols: quercetin 3-O-rutinoside-3′-O-glucoside, quercetin 3,3ʹ-di-O-glucoside, quercetin 3-O-rutinoside, isorhamnetin 3-O-rutinoside, myricetin 3,3ʹ,5ʹ-tri-O-glucoside, myricetin 3,3ʹ-di-O-glucoside and laricitrin 3,5ʹ-di-O-glucoside; and six flavones: luteolin 4ʹ-O-glucoside, apigenin 4′-O-glucoside, tricetin 4ʹ-O-glucoside, tricetin 3ʹ,5ʹ-di-O-glucoside, tricetin 3ʹ-O-glucoside and selagin 5ʹ-O-glucoside, which is a previously undescribed flavone, from their petals. We also compared compositions of floral flavonoid and their aglycone among these species, which suggested that the turquoise species P. alpestris has an essentially intermediate composition between the blue and pale-yellow species. The vacuolar pH was relatively higher in the turquoise (pH 6.2) and pale-yellow (pH 6.2) flower species, while that of blue flower species was usual (pH 5.2). The flower color was reconstructed in vitro using isolated anthocyanin, flavonol and flavone at neutral and acidic pH, and its color was analyzed by reflectance spectra and the visual modeling of their avian pollinators. The modeling demonstrated that the higher pH of the turquoise and pale-yellow species enhances the chromatic contrast and spectral purity. The precise regulation of flower color by flavonoid composition and vacuolar pH may be adapted to the visual perception of their avian pollinator vision.

Bees have been known to visit the male-fertile cultivars of self-incompatible flowering plants more frequently than the male-sterile cultivars, but the origin of this preference is poorly understood. Here, we demonstrate that this preference is driven by the higher protein/lipid ratio of male-fertile pollen compared with male-sterile pollen by way of two caged-behavioral assays with six cultivars. In the first assay, flower-naïve bumblebees (Bombus ignitus Smith) showed a significantly higher flower-visitation rate to male-fertile cultivars (pollen germination rate > 55%; > 14 visits/10 min) of the Japanese pear (Pyrus pyrifolia Nakai) than male-sterile cultivars (pollen germination rate ≤ 20%; > 6 visits/10 min). In the second, bees still preferred the anthers of male-fertile cultivars (5–9 visits/10 min) more than those of male-sterile ones (less than 1 visit in 10 min) even in the absence of all other organs (i.e., petals, pistil, nectar), indicating that pollen is responsible for the preference. We then analyzed the macronutrient content of the pollen and its visual cues, and found that the bee preference was highly correlated with the protein/lipid ratio (0.3–1.6) but not color variables such as (a)chromatic contrast, intensity, and spectral purity. We conclude that the protein/lipid ratio influences the foraging behavior of the bumblebees likely by serving as (1) a chemotactile cue while antennating, (2) a gustatory cue after intake, and (3) an olfactory cue. In addition, the low bee visitation rate to poorly viable pollen could be due to its low protein/lipid ratio.

BACKGROUND: A new physical control method using ultraviolet-B (UV-B) lamps and light-reflecting sheets (UV method) significantly suppressed a spider mite population on greenhouse strawberries. Although UV-B radiation may adversely affect the survival of phytoseiid mites, previous research has suggested that Neoseiulus californicus can improve its survival on exposure to UV-B irradiation by consuming antioxidants contained in tea and peach pollen. In this study, we evaluated strawberry pollen as an alternative food source for N. californicus and examined whether antioxidants in the pollen mitigated UV-B damage to N. californicus. RESULTS: The fecundity of N. californicus females reared on Tetranychus urticae decreased on shifting their diet to pollen. By contrast, females reared continuously on strawberry pollen produced as many eggs as females reared continuously on T. urticae. Survival and fecundity after UV-B irradiation were higher in females on the pollen diet. Oxygen radical absorbance capacity analysis revealed that the high antioxidant activity of strawberry pollen was due to four hydroxycinnamoyl spermidine derivatives. CONCLUSION: Strawberry pollen was an adequate alternative food source for N. californicus. Feeding on strawberry pollen, which contains spermidine derivatives with high antioxidant activity, mitigated UV-B damage. This shows the potential of combining the UV-method with N. californicus for controlling T. urticae in strawberries.

We observed trees of the Japanese apricot, Prunus mume ‘Nanko’ (Rosaceae), bearing two types of flowers: 34% had blue fluorescent pollen under UV irradiation, and 66% had non-fluorescent pollen. The fluorescent pollen grains were abnormally crushed, sterile, and devoid of intine and pollenkitt. The development of microspores within anthers was investigated: in the abnormally developed anthers, tapetal cells were vacuolated at the unicellular microspore stage, and fluorescent pollen was produced. Compounds responsible for the blue fluorescence of pollen were identified as chlorogenic acid and 1-O-feruloyl-β-D-glucose. The anthers with fluorescent pollen contained 6.7-fold higher and 3.8-fold lower amounts of chlorogenic acid and N1,N5,N10-tri-p-coumaroylspermidine, respectively, compared to those with non-fluorescent pollen. The tapetal vacuolization, highly accumulated chlorogenic acid, and deficiency of N1,N5,N10-tri-p-coumaroylspermidine imply that low-temperature stress during the early unicellular microspore stage caused a failure in microsporogenesis. Furthermore, potential effects of the visual difference on the bee behavior were also discussed through the colorimetry. The sterility, likely induced by low-temperature stress, and the preference of honeybees for fluorescence may reduce the pollination efficiency of P. mume.

Most ornithophilous plants have red flowers; this has been associated with ‘the bee avoidance hypothesis’, in which ornithophilous flowers may bear colors that are less conspicuous to bees than melittophilous flowers. In the genus Camellia, C. rusticana and C. japonica bear red flowers and yet recruit different pollinators; the former is entomophilous, while the latter is ornithophilous. C. japonica is considered to have been speciated from a common ancestor later than C. rusticana, accompanying a pollinator shift from insects to birds. Nevertheless, factors explaining the pollinator difference in camellias remain rudimentary. In this study, the color traits of the two camellias were investigated, to determine their color strategy to allure different pollinators. The behavior of bees towards the two camellias was examined by a two-choice assay. Flower color characteristics of the two camellias were analyzed with diffuse reflectance and fluorescence spectra. Based on the visual sensory system of bees and birds, the achromatic contrast, chromatic contrast, intensity, and spectral purity of the two species were evaluated, testing the bee avoidance hypothesis. Furthermore, the compounds responsible for the fluorescence, likely serving as a visual attractant, were identified by NMR and MS. Bees visited C. rusticana flowers almost exclusively and C. japonica hardly at all. Reflectance spectral data showed that C. rusticana petals are more conspicuous to bees than birds due to a UV-reflection secondary peak; and that C. japonica petals exhibited crucially low chromatic contrast against a leaf background to bees, suggesting them to be almost indistinguishable. On the other hand, C. japonica flowers appeared conspicuous to birds. The anthers of C. rusticana exhibited blue fluorescence derived from two anthranilates, while those of C. japonica did not. The two camellias offer different color strategies to be conspicuous to their respective pollinators, and C. japonica seemed to have evolved to avoid bees. Alterations in these color traits may have played a role in pollinator shift.