Fresh Pollen Research Articles

Storage of Bahiagrass Pollen at Different Temperatures

Paspalum is an important genus of grasses, and some species are used for forage in the warmer regions of the world. Efforts to develop improved cultivars often require intra‐ and interspecific hybridization, but asynchronous flowering sometimes prevents hybridization between parental species. One method of circumventing this problem is to store pollen using cryopreservation so pollen will be available to pollinate the female parent when it flowers. Bahiagrass (Paspalum notatum Flügge) was used to determine if Paspalum pollen can be preserved using long‐term cold storage. Pollen was dehydrated with LiCl, MgCl2, NaOH, and silica gel for 30, 60, and 120 min. Both dehydrated and nondehydrated pollen were stored at 4, −20, and −196°C for 10, 60, 120, and 180 d. After thawing, pollen stainability and in vivo germination were used to assess viability. Stainability of fresh pollen was 73.3%, and for treated and stored pollen, it ranged from 25.3 to 71.3%. Storage at −20 and −196°C was superior to storage at 4°C. Pollen dehydrated with LiCl for 30 min stored best at −196°C; pollen dehydrated for 120 min with silica gel and for 30 and 60 min with MgCl2 stored best at −20°C. All samples regardless of treatment, storage temperature, or length of storage had viable pollen. These findings demonstrate that bahiagrass pollen can be stored in long‐term cold conditions, and this procedure can be used to circumvent problems associated with asynchronous flowering between parental species used in wide hybridization programs.

Pollen has higher water content when dispersed in a tricellular state than in a bicellular state

ABSTRACT Pollen is generally dispersed in a sexually immature and somewhat dehydrated, metabolically quiescent state. Yet, in some species, pollen at anthesis is well-hydrated and metabolically active, and in 30 % of angiosperms pollen is dispersed after having formed its sperm cells. Pollen water content and sexual maturity may be correlated, either because both are subject to trade-offs between dispersal viability and post-pollination performance, or because the traits display developmental linkages. We inferred relative water content of sexually immature (“bicellular”) and sexually mature (“tricellular”) pollen of 30 species of angiosperms using a hydration index (HI) that ranges from zero to one, based on how near fresh pollen volume is to its minimal (dehydrated) or maximal (hydrated) volume. Tricellular pollen had 30 % higher HI than bicellular pollen (P < 0.005), after controlling for initial pollen size (larger pollen had higher HI; P < 0.05). A literature survey of 344 species indicated that the tricellular and hydrated states were strongly associated, although all four trait state combinations were present (P < 0.0001). Our results suggest that a common mechanism for the repeated origins of tricellular pollen has been via the loss of controlled pollen dehydration, which enables either accelerated or extended pollen development.

Pesticide residues in honey bees, pollen and beeswax: Assessing beehive exposure

In order to study the distribution of pesticide residues in beekeeping matrices, samples of live in-hive worker honey bees (Apis mellifera), fresh stored pollen and beeswax were collected during 2016–2017 from 45 apiaries located in different landscape contexts in Spain. A total of 133 samples were screened for 63 pesticides or their degradation products to estimate the pesticide exposure to honey bee health through the calculation of the hazard quotient (HQ). The influence of the surrounding environment on the content of pesticides in pollen was assessed by comparing the concentrations of pesticide residues found in apiaries from intensive farming landscapes to those found in apiaries located in mountainous, grassland and urban contexts. Beeswax revealed high levels of miticides used in beekeeping such as coumaphos, chlorfenvinphos, fluvalinate and acrinathrin, which were detected in more than 75% of samples. Pollen was predominantly contaminated by miticides but also by insecticides used in agriculture such as chlorpyrifos and acetamiprid, which showed concentrations significantly higher in apiaries located in intensive farming contexts. Pesticides residues were less frequent and at lower concentrations in live honey bees. Beeswax showed the highest average hazard scores (HQ > 5000) to honey bees. Pollen samples contained the largest number of pesticide residues and relevant hazard (HQ > 50) to bees. Acrinathrin was the most important contributor to the hazard quotient scores in wax and pollen samples. The contributions of the pesticides dimethoate and chlorpyrifos to HQ were considered relevant in samples.

Staining for viability testing, germination and maturation of Sambucus nigra L. pollen in vitro

Freshly released pollen of black elderberry (Sambucus nigra L.) was incubated under various culture conditions until germination was achieved. Optimal conditions for germination were determined and used for maturation of unicellular microspores in vitro. Staining with 5-diphenyltetrazolium bromide, propidium iodide and iodine potassium iodide was used to assess pollen viability, nuclear phase and maturation, respectively. The germination rate was highest when fresh pollen was agitated at 40 rpm in Petri dishes containing a liquid medium consisting of Brewbaker and Kwack salts, 15% (w/v) sucrose, 500 mg/l MES sodium salt, at pH 5.0; germination reached nearly 70% after only 1 h in culture. Under these conditions, and with addition of 200 mg/l glutamine, 260 mg/l cytidine and 500 mg/l uridine, uninucleate microspores developed into mature pollen at a 12% germination rate. Our report is the first demonstration of maturation of S. nigra microspores in vitro.

Physico-chemical, microbiological, and antioxidant activity of fresh bee pollen, dried bee pollen, and bee pollen powder of Apis mellifera

Physico-chemical, microbiological, and antioxidant activity of fresh bee pollen, dried bee pollen, and bee pollen powder of <i>Apis mellifera</i>

Suitability of date palm pollen as an alternative food source for the predatory mite Cydnoseius negevi (Swirski and Amitai) (Acari: Phytoseiidae) at a low relative humidity

The nutritional suitability of date palm pollen for the predatory mite Cydnoseius negevi (Swirski and Amitai) (Acari: Phytoseiidae) was tested by assessing the development and life table parameters of the predator at two relative humidity levels, 35 and 60 % RH, at 30 °C in the laboratory. C. negevi successfully developed and reproduced when fed on a fresh date palm pollen diet at both humidity levels. However, at 35 % RH, the incubation period and development of immatures were significantly longer, female longevity and fecundity significantly decreased, and all life table parameter values were negatively affected. The results obtained in this study were compared with those of some previously published studies in the same field. The date palm pollen could be a suitable food source for the mass production of C. negevi. Moreover, the successful reproduction and development of C. negevi fed date palm pollen at low RH (35 %) makes it a good alternate food source in the field to conserve the population of the predator under local arid environmental conditions.

BACTERIAL CONTAMINATION OF VACUUM STORED FLOWER BEE POLLEN

Water content, water activity (Аw) and mean microbiological characteristics were determined in fresh and dried flower bee pollen collected from apiary from North east region in Bulgaria. Higher microbial counts were established for fresh than for dried bee pollen after long-time (1 and 2-years) vacuum-packed cold storage (0-4 ºС and - 18 ºС). It was found contamination with Pantoea spp., Prot. mirabilis, Citrob. freundii, Serratia fonticola, Raoultella planticola/ornithinolytica, S. hominis (subspecies hominis), S. sciuri (subspecies sciuri), Flavimonas orizihabitans, Achromobacter denitrificans, Arthrobacter globiformis, Psychrobacter phenylpyruvicus, B. pumilis and Enterococcus faecalis in the dried flower bee pollen intended for human consumption. Fresh bee pollen that is an intermediate product not intended for human consumption besides these contained Prot.vulgaris, E.coli, Staph. epidermidis and B. subtilis.

CAT and MDH improve the germination and alleviate the oxidative stress of cryopreserved Paeonia and Magnolia pollen

Researches have reported that reactive oxygen species (ROS)-induced oxidative stress plays an important role in cell cryodamage during cryopreservation. In the current study, pollen from Magnolia denudata and Paeonia lactiflora ‘Zi Feng Chao Yang’ was cryopreserved and incubated with exogenous catalase (CAT) and malate dehydrogenase (MDH) immediately after thawing. The effect of CAT and MDH on the germination of cryopreserved pollen was measured. Based on that, the ROS level, lipid peroxidation and antioxidants activities in fresh pollen, cryopreserved pollen added with or without CAT or MDH were determined to investigate their relationship with oxidative stress. Pollen from Magnolia and Paeonia showed a significant loss of germination, but marked increase of ROS and malondialdehyde (MDA) production after cryostorage. Antioxidant profiles in them were also enhanced. CAT and MDH addition increased the post-LN pollen germination of Magnolia and Paeonia significantly. Their germination rate achieved the highest with 100 IU ml−1 MDH and 400 IU ml−1 CAT application, respectively. Compared to their untreated controls, ROS and MDA accumulation reduced significantly in cryopreserved Magnolia pollen treated with 100 IU ml−1 MDH, while superoxide dismutase (SOD) activity improved markedly. In the case of Paeonia, significantly lower level of ROS and MDA, but higher activity of CAT and SOD were observed in cryopreserved pollen treated with 400 IU ml−1 CAT. In conclusion, pollen deterioration after cryopreservation is associated with ROS-induced oxidative stress. Exogenous CAT and MDH can reduce the oxidative damage through the activity stimulation of antioxidant enzymes, and play a protective role in the pollen during cryopreservation.

Efficacy of sonication-assisted sieving on Quaternary pollen samples

ABSTRACTDebris in archaeological pollen samples can obscure pollen grains during microscopic analysis. When attempting to remove debris smaller than 10 microns from a pollen sample, screen openings can be too small, requiring the sample to be agitated in order to facilitate its passing through the screen. We recently developed a new method that uses a Branson© S450 sonicating disruptor horn to agitate the sample, keeping the screen free from debris. Using this method, we have been successful in ridding the pollen sample of debris smaller than 10 microns. This paper presents the results of an investigation of the potential effects of sonication on pollen preservation. We selected 12 fresh pollen types based on a unique range of morphological characteristics, and subjected this assemblage to sonication for different lengths of time and at variable output intensities. Our study demonstrated that when used for 3 minutes at low to medium output intensities, a sonicating disruptor horn can aid in the removal of unwanted debris without considerably damaging pollen grains. We also did not find evidence that pollen colliding with other debris in a sample increased levels of degradation. Analysis on the efficacy of sonication-assisted sieving demonstrated that more tiny debris was removed from the sample at higher frequencies and over increasing durations.

Laboratory rearing and food consumption of spring bumble bee (Bombus haemorrhoidalis Smith) queens

Spring collected bumble bee queens were reared under laboratory condition at 27 ± 1°C temperature and 65–70% relative humidity and by continuously feeding the measured quantity of sucrose solution (50%) and fresh pollen pellets collected by honeybees. The average duration for wax secretion was 14.27 ± 1.99 days, whereas average development time from egg to adult in workers of B. haemorrhoidalis was observed to be 24 ± 1.1 days. Queen consumed 1.77 ml sucrose and 0.78 g pollen daily.

Digestibility and nutritional value of fresh and stored pollen for honey bees (Apis mellifera scutellata)

Pollen, the main protein source for honey bees, is mixed with regurgitated nectar or honey during collection and then stored as ‘bee bread’ before its consumption, mainly by young nurse workers. It has been suggested that storage of pollen improves its nutritional value and digestibility, but there is little evidence for such changes. We fed two fresh pollen types of different protein content (aloe and sunflower), and two stored pollen types (sunflower and a mixed pollen), to young caged worker bees. We measured daily consumption of pollen and sucrose solution, and survival after 14 days. At day 14 we recorded ovarian activation and extraction efficiency, by counting empty pollen grains in the rectal contents. Extraction efficiency is a measure of pollen digestibility. Contrary to our predictions, bees did not consume more fresh sunflower pollen than fresh aloe pollen to compensate for the lower protein content of sunflower pollen. In addition, they did not consume less sucrose solution when fed stored pollen diets that are already enriched in sugar. Consumption of stored sunflower pollen resulted in a low protein to carbohydrate (P:C) intake. Survival and ovarian activation were higher on diets giving higher P:C intakes. Extraction efficiency was high (up to 99%) for all pollen diets, and comparison of fresh and stored sunflower pollen showed that storage did not make it easier to digest. Changes to pollen during storage do not confer obvious benefits to honey bees.

Chemical characterization and identification of Pinaceae pollen by infrared microspectroscopy.

FTIR microspectroscopy, in combination with spectral averaging procedure, enables precise analysis of pollen grains for chemical characterization and identification studies of fresh and fossilised pollen in botany, ecology and palaeosciences. Infrared microspectroscopy (µFTIR) of Pinaceae pollen can provide valuable information on plant phenology, ecophysiology and paleoecology, but measurements are challenging, resulting in unreproducible spectra. The comparative analysis of µFTIR spectra belonging to morphologically different Pinaceae pollen, namely bisaccate Pinus and monosaccate Tsuga pollen, was conducted. The study shows that the main cause of spectral variability is non-radial symmetry of bisaccate pollen grains, while additional variation is caused by Mie scattering. Averaging over relatively small number of single pollen grain spectra (approx. 5-10) results with reproducible data on pollen chemical composition. The practical applicability of the µFTIR spectral averaging method has been demonstrated by the partial least-squares regression-based differentiation of the two closely related Pinus species with morphologically indistinguishable pollen: Pinus mugo (mountain pine) and Pinus sylvestris (Scots pine). The study has demonstrated that the µFTIR approach can be used for identification, differentiation and chemical characterization of pollen with complex morphology. The methodology enables analysis of fresh pollen, as well as fossil pollen from sediment core samples, and can be used in botany, ecology and paleoecology for study of biotic and abiotic effects on plants.

Low Temperature Storage and in vitro Pollen Germination of Selected Spring Wheat Accessions

The study was conducted to evaluate in vitro pollen germination of 50 spring wheat (Triticum aestivum L.) accessions at three storage temperatures (23 °C, 5 °C, and -20 °C). Germination of the mature pollen was measured right after harvest (0 time), and 24, 48, and 72 h after storage (HAS). Differences in fresh pollen germination between accessions were significant and ranged from 21.15% to 1.09%. Pollen germination was the lowest at 24 h at room temperature. The pollens of 24 wheat accessions (AAC Scotia, AW636, AW725, AW780, AW804, AW822, AW823, Brookfield, BRS Pardela, ECSW05, ECSW38, ECSW48, ECSW49, ECSW69, Hoffman, Kleos, Major, Nass, 12NQW-237, 12NQW-413, 12NQW-414, 12NQW-436, 12NQW-754, and 12BW0374) were viable after 24 h when they were stored at 5 °C. Sub-zero temperature did not prolong pollen viability.

ROS-induced oxidative stress is closely related to pollen deterioration following cryopreservation

Oxidative stress induced by excessive accumulation of reactive oxidative species (ROS) during cryopreservation is thought to be one factor contributing to cryodamage of biological materials. To explore the role of oxidative stress in the cryopreservation of plant pollen, germination, ROS, and malondialdehyde (MDA) levels of pollen from 20 ornamental plant species were compared before and after cryopreservation. The results showed that the germinability of cryopreserved pollen from 13 out of the 20 species was not significantly different from that of fresh pollen (group 1), only one increased significantly, while the other six declined significantly (group 2). The MDA content in cryopreserved pollen from nine species in group 1 showed no significant difference from that of fresh pollen, while four species in group 2 rose significantly. This suggested that pollen viability and MDA levels were negatively correlated. ROS generation in cryopreserved pollen from nine species in group 1 was unchanged compared to fresh pollen, while five species in group 2 increased significantly. This suggested that pollen viability was negatively correlated with ROS generation. Additionally, both ROS and MDA levels in pollen from four species in group 2 increased significantly. In conclusion, pollen from most species possesses some cryostorage tolerance, but some species are severely damaged by cryostorage. Oxidative stress induced by the cryostorage in liquid nitrogen (LN) may be a key factor for the decreased viability in pollen following cryopreservation.

Revealing Pesticide Residues Under High Pesticide Stress in Taiwan's Agricultural Environment Probed by Fresh Honey Bee (Hymenoptera: Apidae) Pollen.

Significant pesticide residues are among the most serious problems for sustainable agriculture. In the beekeeping environment, pesticides not only impact a honey bee's survival, but they also contaminate bee products. Taiwan's agricultural environment has suffered from pesticide stress that was higher than that found in Europe and America. This study deciphered problems of pesticide residues in fresh honey bee pollen samples collected from 14 monitoring apiaries in Taiwan, which reflected significant contaminations within the honey bee population. In total, 155 pollen samples were screened for 232 pesticides, and 56 pesticides were detected. Among the residues, fluvalinate and chlorpyrifos showed the highest concentrations, followed by carbendazim, carbaryl, chlorfenapyr, imidacloprid, ethion, and flufenoxuron. The average frequency of pesticide residues detected in pollen samples was ca. 74.8%. The amounts and types of pesticides were higher in winter and in southwestern Taiwan. Moreover, five of these pollen samples were contaminated with 11-15 pesticides, with average levels between 1,560 and 6,390 μg/kg. Compared with the literature, this study emphasized that pollen gathered by honey bee was highly contaminated with more pesticides in Taiwan than in the America, France, and Spain. The ubiquity of pesticides in the pollen samples was likely due to the field applications of common pesticides. Recently, the Taiwanese government began to improve the pesticide policy. According to the resurvey data in 2016, there were reductions in several pesticide contamination parameters in pollen samples from west to southwest Taiwan. A long-term investigation of pollen pesticide residues should be conducted to inspect pesticides usage in Taiwan's agriculture.

Pollinic characterization of Raulinoa echinata R. S. Cowan (Rutaceae), Dyckia brevifolia Baker and Dyckia ibiramensis Reitz (Bromeliaceae), reophyte and saxicolous endemic species of river Itajaí-Açu, Santa Catarina, Brazil

Aiming to contribute with elements about the reproduction of the reophytes Raulinoa echinata, Dyckia brevifolia and Dyckia ibiramensis as well as to support their taxonomic definition, their pollinic characterization was carried out. Fresh pollen grains were prepared by acetolysis. Observations occurred on a light microscope and a scanning electron microscope. Measures were done of the polar (P) and equatorial diameters (E) and thickness of the exine and are expressed as means in micrometers. R. echinata: monad; radial symmetry; medium size; P=49,07±3,33; E=33,95±3,94; prolate (ratio P / E); ambitus subtriangular; 3-aperturated with long colpi and lalongate endoapertures; exine striated with thickness 1.54 ± 0,32. Dyckia brevifolia: monad; bilateral symmetry; medium size; P=25,16 ± 3,02; E=43,16±3,50 x 28,12±3,26; ambitus elliptical; monoaperturated with one long sulcus; exine reticulate with thickness 1,66 ± 0,31. D. ibiramensis: monad; bilateral symmetry; medium size; P=29,54±3,17; E=42,36±3,60 x 28,43±3,38; ambitus elliptical; monoaperturated with one long sulcus; exine reticulate with thickness 1,73 ± 0,28. Data obtained can allow to verify hydrophily for the three species as well as suggest Raulinoa proximity with Metrodorea and Esenbeckia (both Pilocarpinae) and confirm D. brevifolia e D. ibiramensis to Pitcairnioideae.

Pollinic characterization of Raulinoa echinata R. S. Cowan (Rutaceae), Dyckia brevifolia Baker and Dyckia ibiramensis Reitz (Bromeliaceae), reophyte and saxicolous endemic species of river Itajaí-Açu, Santa Catarina, Brazil

Aiming to contribute with elements about the reproduction of the reophytes Raulinoa echinata, Dyckia brevifolia and Dyckia ibiramensis as well as to support their taxonomic definition, their pollinic characterization was carried out. Fresh pollen grains were prepared by acetolysis. Observations occurred on a light microscope and a scanning electron microscope. Measures were done of the polar (P) and equatorial diameters (E) and thickness of the exine and are expressed as means in micrometers. R. echinata: monad; radial symmetry; medium size; P=49,07±3,33; E=33,95±3,94; prolate (ratio P / E); ambitus subtriangular; 3-aperturated with long colpi and lalongate endoapertures; exine striated with thickness 1.54 ± 0,32. Dyckia brevifolia: monad; bilateral symmetry; medium size; P=25,16 ± 3,02; E=43,16±3,50 x 28,12±3,26; ambitus elliptical; monoaperturated with one long sulcus; exine reticulate with thickness 1,66 ± 0,31. D. ibiramensis: monad; bilateral symmetry; medium size; P=29,54±3,17; E=42,36±3,60 x 28,43±3,38; ambitus elliptical; monoaperturated with one long sulcus; exine reticulate with thickness 1,73 ± 0,28. Data obtained can allow to verify hydrophily for the three species as well as suggest Raulinoa proximity with Metrodorea and Esenbeckia (both Pilocarpinae) and confirm D. brevifolia e D. ibiramensis to Pitcairnioideae.

Water activity of fresh bee pollen and mixtures of bee pollen-honey of different botanical origin

A mixture of bee pollen and honey could represent a complete food supplement for human diet. In this study, for three bee pollen-honey mixtures ratios at two storage times (0 and 90 days) at room temperature, water activity (aw) were investigated. For the mixtures were employed Castanea and Eucalyptus pollen species, and acacia, chestnut and “beach” honeys. In the mixtures, Castanea and Eucalyptus pollen showed a different aw trend in relation to pollen concentration and to storage time. In Castanea-honeys mixtures and in Eucalyptus-honey mixtures (only at time 0) aw values were higher than each component of the corresponding mixtures. In Eucalyptus-honeys mixtures (only at time 90) aw values were lower than each component. This investigation highlights that honey, bee-pollen and their mixtures, stored at room temperature, could have a little change in their aw content, in time. Results shows that aw values of bee pollen mixed with honey were all lower than 0.7, although bee pollen-honey mixtures were stored at room temperature instead at freezing temperature (−15 °C) for 90 days.

Quantitative Analysis of Long-Term Mass Rearing of Neoseiulus californicus (Acari: Phytoseiidae) on Almond Pollen.

Long-term feeding effects of the almond pollen on the life table parameters of Neoseiulus californicus McGregor were assessed after 5, 10, and 20 generations after introduction in the rearing arena. Furthermore, to evaluate behavioral characteristics of the mass-reared predator (strain A) in face with the real prey, functional and numerical responses of the predator to different densities of the twospotted spider mite nymphs were determined, and the obtained data were compared with those reared on twospotted spider mite (strain T). Long-term rearing did not significantly affect total fecundity of N. californicus (ranged from 37.79 to 41.91 eggs). Nevertheless, preadult duration in the 5th generation was significantly longer than the 10th and 20th generations. The intrinsic rate of increase (r) in the 10th (0.2056 d-1) and 20th (0.2201 d-1) generations had not significant difference together. However, the r value slightly dropped in the 5th generation (0.1706 d-1) because of the irregular offering of fresh pollen to the rearing colonies before that. Both strains of N. californicus exhibited a type II functional response; however, the N. californicus reared on the almond pollen (strain A) had a higher attack rate (a) and shorter handling time (Th). The individuals reared on the almond pollen had a greater size than those reared on twospotted spider mite; its higher predation potential is probably due to this characteristic. Consequently, the rearing of N. californicus on the almond pollen positively affected its attributes including high survivorship, body size, and fecundity, and subsequently higher potential to control twospotted spider mite.

Pollen drying and storage for hybrid seed production of cucumber (Cucumis sativus L.)

One of the major problems in hybrid seed production of cucumbers is the low pollen production and low pollen viability. This research was mainly dealt with pollen handling in order to maintain its viability during storage, which was carried out in three steps. Male parental of 'KE014' was planted at Production Farm of East West Seed Indonesia Ltd. in Jember, East Java, Indonesia during March to September 2012. In a first step, male flowers were harvested and pollen was extracted and dried for 24 h during which the water content and pollen viability was monitored. In a second step the dried pollen was stored in an ultra-freezer (-79±2°C) for 56 days and its viability was also monitored every 7 days. The third step was the pollination of a female parental stock to produce hybrid seed. The experiments were arranged in three replications with completely randomized design. Results showed that pollen water content decreased significantly during the first 8 h of drying from 12.05 to 6.71% while the pollen viability increased from 10.16 to 25.60%. Prolonged pollen drying time up to 24 h did not substantially reduce the water content. Pollen dried for 8 h was able to maintain its viability during 56 days of storage in ultra-freezer (-79±2°C) better than those undried or dried longer. The fruit set and seed set got from pollination with undried and stored pollen (11.0 and 23.0%, respectively) was lower than fruit set and seed set got with dried and stored pollen (ranged 46.5-58.9 and 52.0-67.8%, respectively). Using stored pollen (undried or dried) for pollination in hybrid seed production yielded in lower number of filled seed per fruit than using fresh pollen. Hybrid seed germination got from pollination with dried and stored pollen was slightly higher (98.36-99.63%) than those got from undried and stored pollen (95.58%), nevertheless all of them were high quality seeds. Thus, using the stored pollen for pollination in cucumber hybrid seed production would reduce seed yield but not seed quality.