FOOD PLANTS OF THE RED MASON BEE (Osmia rufa L.) DETERMINED BASED ON A PALYNOLOGICAL ANALYSIS OF FAECES

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1 Vol. 51 No Journal of Apicultural Science 55 FOOD PLANTS OF THE RED MASON BEE (Osmia rufa L.) DETERMINED BASED ON A PALYNOLOGICAL ANALYSIS OF FAECES Dariusz Teper Research Institute of Pomology and Floriculture, Apiculture Division. ul. Kazimierska 2, Pu³awy, Poland. dariusz.teper@man.pulawy.pl Received 15 September 2007; accepted 12 November 200 S u m m a r y Samples of faeces to be analyzed were collected in the years To this end, each day after dark over the entire flight period of red mason bees, their colonies were covered, from the side of tube outlets, with an isolator made of a fine screen. The next day, while trying to exit, the bees swarmed the isolator and excreted faeces onto its surface. Microscopic preparations were made that were then subjected to a palynological analysis. In the two years of the experiments, a total of 56 samples of faeces were collected, in which 34 types of pollen of plants were identified belonging to 22 families. Most of the identified types of pollen originated from nectariferous species, yet as many as 29% (10 species) belonged to taxa providing insects mainly with pollen. They also included anemophilous trees Fagus, Juglans, Pinus, Quercus as well as weeds Plantago and Rumex. A small part of the pollen was not possible to be determined due to the considerable extent of its degradation in the process of digestion. Based on the results of a palynological analysis, a food supply sequence of Osmia rufa was prepared for each experimental year. Keywords: Osmia rufa, food plants, pollen analysis of faeces, digestion of pollen. INTRODUCTION For Polish crops, the main pollinating insect is the honeybee, which forms large, solid and all-season avilable colonies. Under climatic conditions of Poland, the highest demand for pollinating insects occurs in the last decade of April and in May when the number of wild Apidae insects is small since bumble bee colonies are then at the initial stage of development, whereas most of species of solitary bees begin their flights later due to an elongated developmental cycle. The red mason bee (Osmia rufa) belongs to a small number of species wintering in the imago stage and leaving cocoons as early as April; its flight season spans till June (Ruszkowski et al 1998). Natural colonies of Osmia rufa are not numerous due to a lack of appropriate sites for nesting and to parasites reducing their population. Owing to its relatively undemanding culture (Wójtowski and Wilkaniec 1978) it is possible to reproduce colonies to considerable sizes and to use those bees for pollination of also large commercial plantations. Research carried out for a number of years have proved the high usability of Osmia rufa as a pollinator of entomophilous plants. Results of investigations to date have been based on time-consuming observations of bee flights (Ruszkowski and Biliñski 1986) or on a palynological analysis of pollen accumulated by Osmia females in nests (Wilkaniec and Warakomska 1992, Wilkaniec et al 1997, Biliñski and Teper 2004). In the last case, the collection of pollen samples was linked with damage to eggs or larvae of progeny generation. In my study, food plants of Osmia rufa were identified using a non-invasive method of pollen analysis of bee faeces. It

2 56 was possible due to the fact that pollen is incompletely digested by the insects, and that bee faeces contain its residues often in an unaltered, very resistant form of outer wall built of sporopollenins exine. This study was aimed at verifying whether the method of a pollen analysis of faeces, being safer to insects and less labor-consuming than the other observational methods, may be applicable in the identification of plants visited by Osmia rufa. MATERIALS AND METHODS In the experiment carried out in the years , use was made each year of over 1,000 cocoons of Osmia rufa L. originating from the culture reared for several years at the Apiculture Division, Institute of Pomology and Floriculture in Pu³awy. Colonies of solitary bees were located in Pu³awy in an area adjacent to home gardens and rapeseed plantations. Samples of faeces were collected from the 8 th of May till the 6 th of June 2003 and between the 15 th of May and 12 th of June Collection of samples of Osmia rufa faeces Due to the colonial character of life of solitary bees, the method of their faeces sample collection had to be elaborated that differed compared to the method used for bumble bees (Teper 2006). Use was made of the fact that red mason bees spend nights in self-established nests. In a box chest (35x40 cm) ca. 1,000 of reed tubes with internal diameters of 5 12 mm and length of cm were fixed. A cardboard box with slots containing 1,000 cocoons of Osmia rufa was hung under the nest. Once the females were brought out and began establishing nests, each day at dusk, upon termination of bee flights, an isolator made of chiffon was put onto on the box chest and a girt with a rubber band so that it adhered tightly to the walls of the box chest (Photo 1). Outlets of nest tubes were directed to the East, owing to which the bees began their flight early in the morning. Trying to leave the nest, the bees swarmed the isolator screen and excreted faeces onto its surface. Usually, at around 9:00. a.m. the isolator was taken off and the bees were let out. Sometimes, on chilly or cloudy days, the colony was left isolated for 1-2 hours longer. Photo 1. Isolated colony of Osmia rufa The isolator covered with faeces was dried, and the faeces were later disintegrated onto white paper and poured into specifically designed and tagged paper bags, in which the faeces were dried in a dry state in a refrigerator. Samples were collected each day over the entire flying period of bees except for rainy days when precipitation washed the faeces off the isolator. Execution of microscopic preparations from faeces of Osmia rufa On the termination of Osmia rufa flights, faeces stored in paper bags in a refrigerator were poured into pits of plates for spot tests and 1-2 drops of distilled water were later added to each of them and the plates were left for a few hours for soaking (Photo 2). Afterwards, the resultant suspensions were thoroughly stirred by means of a glass rod, and smears were performed with an inoculation loop onto tagged microscopic slides. Slightly pre-dried preparations were covered with microscopic cover

3 Vol. 51 No Journal of Apicultural Science 57 Photo 2. Samples of faeces of Osmia rufa in pits of spot plates glasses with the addition of glycerol gelatin. Microscopic analysis of the collected material Pollen grains of microscopic preparations were identified with the use of available pollen atlases (among others Sawyer 1981, 1988; Ricciardelli d`albore 1998; Bucher et al. 2004) and own collection of over 300 reference preparations using the classification by Zander (1935, 1937, 1941, 1949, 1951). In each microscopic preparation pollen was determined, when possible, into genus, species, family or type of build and described in a table. RESULTS Over the two experimental years, a total of 56 microscopic preparations were made in which 34 types of pollen were identified (belonging two 22 families): Acer, Aesculus, Anthriscus type, Asteraceae, Brassicaceae, Caryophyllaceae, Clematis, Fagus, Filipendula, Frangula, Juglans, Lilium, Lonicera, Lotus, Lythrum, Melilotus, Pinus, Plantago, Potentilla type, Prunus type, Pterocarya, Quercus, Ranunculus, Robinia, Rosaceae, Rubus type, Rumex, Symphytum, Taraxacum type, Tilia, Trifolium repens s.l., Viola, Viscum album, and Viburnum. In faeces of 2003 there were identified 28 types of pollen (Table 1), whereas in those of types (Table 2). Most of them were constituted by species providing nectar and pollen, yet as many as 29% (10 species) belonged to taxa providing insects mainly with pollen. They also included anemophilous trees Fagus, Juglans, Pinus, and Quercus as well as weeds Plantago and Rumex. In both experimental years, flights of Osmia rufa lasted for ca. one month. Some small part of pollen could not be determined due to considerable damage of exine during the digestion process. The damaged grains belonged, probably, to species of plants secreting pollen with the thickest exine and the most susceptible to conditions occurring in the gastrointestinal tract of insects. Detailed analysis of samples of Osmia rufa faeces enabled determining a food supply sequence for each experimental year (Table 1, 2), according to the method of Ruszkowski and ak (1974), depicting changes in the species composition of visited plants over the flying season. DISCUSSION The pollen analysis of faeces collected from colonies of Osmia rufa in the years 2002 and 2004 enabled the identification of 34 types of pollen of visited plants. However, it constitutes merely 30% of the number of species reported by Ruszkowski and Biliñski (1986). A substantially number of taxons enumerated by those authors is certainly linked with vast material collected in the years in the area of the entire country. Those observations covered exclusively entomophilous plants, hence a lack of any notice on anemophilous plants, especially trees that constitute a very important source of pollen to Osmia rufa larvae. The fact that those plants had been visited was proven by the pollen analysis of faeces which were often found to contain pollen of Quercus, Juglans or Pinus. This was also confirmed by studies of plants visited by Osmia based on the

4 58 pollen analysis of the contents of nesting cells conducted by Wilkaniec and Warakomska (1992, 1997) as well as by Biliñski and Teper (2004). The special interest of Osmia rufa in visiting the anemophilous plants, exceeding multiple the pollen yield of entemophilous plants, may be explained by vast demand of insects for pollen indispensable for feeding larvae (in each cell, females store ca. 200 mg of pollen). The high preferences of the Osmia rufa for anemophilous plants are likely to be the reason of lower interest in visiting entemophilous crops blooming in the same period. The presented food supply sequence of Osmia elaborated based on the pollen analysis of their faeces has no equivalent in literature data. Another, very important aspect is the fact that no single nesting cell of Osmia was damaged during elaboration of the list of plants visited by the insects by means of a pollen analysis of their faeces, whereas in the previous study several dozen nests containing a total of a few hundred progeny cells were destroyed in each analysis. A negligible (since estimated at a level of ca. 10%) number of types of pollen grains that were not able to be identified in this study does not affect the practical applicability of the method.

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7 Vol. 51 No Journal of Apicultural Science 61 CONCLUSIONS In a two-year study carried out on plants visited by Osmia rufa, the pollen analysis of their faeces enabled determining 34 taxa of visited plants. The frequent presence of pollen of anemophilous plants in faeces of Osmia rufa often indicates its feeding preferences towards these taxa. The presented analytical method may be applied in studies of host plants of other sparse, protected species of insects, as it does not require their catching or destruction of their nests. REFERENCES Anasiewicz A. (1992) Pokarm roœlinny biedronki siedmiokropki (Coccinella septempunctata). XLI Zjazd Pol. Tow. Entomol., Wroc³aw. Biliñski M., Teper D. (2004) Chów i wykorzystanie murarki ogrodowej Osmia rufa (Hymenoptera, Megachilidae) do zapylania sadów. J. Apic. Sci. 48 (2): Brian A.D. (1951) The pollen collected by bumble-bees. J. Anim. Ecol. 20: Bucher E., Kofler V., Vorwohl G., Zieger E. (2004) Das Pollenbild der Südtirolen Honige. Biologisches Labor der Landesagentur für Umwelt und Arbeitsschutz. Golding Y., Edmunds M. (2003) A novel method to investigate the pollen diets of hoverflies. Journal of Botanical Education, 37(4): Koz³owska A., Warakomska Z. (1984) Biologia i znaczenie niektórych po ytecznych muchówek. Wszechœwiat, 85 (11): Ricciardelli d`albore G. (1998) Mediterranean melissopalynology. University of Perugia, Italy, pp.466. Ruszkowski A., Biliñski M. (1986) Roœliny pokarmowe oraz znaczenie gospodarcze murarek. Pszczeln. Zesz. Nauk. 30: Ruszkowski A. Gosek J., Biliñski M., Pawlikowski T., Kosior A., Fija³ J., Kaczmarska K. (1998) Okresy pojawu pszczó³ samotnic z rodziny miesiarkowatych (Hymenoptera, Apoidea, Megachilidae) oraz przebieg inkubacji niektórych gatunków. Pszczeln. Zesz. Nauk. 42 (1): Ruszkowski A., ak B. (1974) Taœma pokarmowa wa niejszych gatunków trzmieli (Bombus Latr.) oraz mo liwoœci ich rozmna ania. Pamiêtnik Pu³awski, supl. do z. 58: Sawyer R. (1981) Pollen Identification for Beekeepers. University College Cardiff Press, UK, pp Sawyer R. (1988) Honey identification. Cardiff Academic Press, UK, pp Teper D. (2003) Mo liwoœæ okreœlania gatunków roœlin oblatywanych przez trzmiele na podstawie analizy palinologicznej ich odchodów. Materia³y z XL Naukowej Konferencji Pszczelarskiej, Pu³awy marca 2003 r.: Teper D. (2004) Food plants of Bombus terrestris L. determined by palynological analysis of pollen loads. J. Apic. Sci., 48 (2): Wilkaniec Z., Warakomska Z. (1992) Host plants of Osmia rufa L. defined on the basis of pollen stored by female bees. In: Natural resources of wild bees in pollen. Pedagogical University, Bydgoszcz: Wilkaniec Z., Warakomska Z., Giejdasz K. (1997) Roœliny pokarmowe populacji Osmia rufa L. (Apoidae, Megachilidae) zlokalizowanej w wielkotowarowym gospodarstwie Swadzim. Postêpy apidologii w Polsce. Wyd. Uczelniane WSP w Bydgoszczy: Wójtowski F., Wilkaniec Z. (1978) Hodowla i u ytkowanie pszczó³ samotnic osiedlonych w pu³apkach gniazdowych. Instr. Wdro. AR, Poznañ, p.10. Zander E. (1935, 1937, 1941, 1949, 1951) Beiträge zur Herkunftsbbestimmung bei Honig. I Reichsfachgruppe Imker, Berlin; II, III, V Liedloff Loth & Michaelis, Leipzig; IV Ehrenwirth, München: 1100 pp.

8 62 ROŒLINY POKARMOWE MURARKI OGRODOWEJ (Osmia rufa L.) OKREŒLANE NA PODSTAWIE ANALIZY PALINOLOGICZNEJ ODCHODÓW Teper D. S t r e s z c z e n i e Próbki odchodów do badañ pobierano w latach W tym celu codziennie po zmroku przez ca³y okres lotu murarek kolonie os³aniano, od strony wylotów z rurek, izolatorem z gêstej siatki. Nastêpnego dnia rano pszczo³y chc¹c siê wydostaæ na zewn¹trz obsiada³y izolator i wydala³y odchody na jego powierzchniê. Z odchodów wykonywano preparaty mikroskopowe, które nastêpnie poddano analizie palinologicznej. ¹cznie w ci¹gu dwóch lat badañ pobrano 56 próbek odchodów. Zidentyfikowano w nich 34 typy py³ku roœlin nale ¹cych do 22 rodzin: Acer, Aesculus, typ Anthriscus, Asteraceae, Brassicaceae, Caryophyllaceae, Clematis, Fagus, Filipendula, Frangula, Juglans, Lilium, Lonicera, Lotus, Lythrum, Melilotus, Pinus, Plantago, typ Potentilla, typ Prunus, Pterocarya, Quercus, Ranunculus, Robinia, Rosaceae, typ Rubus, Rumex, Symphytum, typ Taraxacum, Tilia, Trifolium repens s.l., Viola, Viscum album, Viburnum. W odchodach z 2003 roku oznaczono 28 typów py³ku (Tab. 1), a w 2004 roku 22 typy (Tab. 2). Wiêkszoœæ oznaczonych typów py³ku pochodzi³a z gatunków nektarodajnych, jednak a 29% (10 gatunków) nale a³o do taksonów dostarczaj¹cych owadom g³ównie py³ku. By³y wœród nich równie wiatropylne drzewa buk (Fagus), orzech (Juglans), sosna (Pinus), d¹b (Quercus) oraz chwasty babka (Plantago) i szczaw (Rumex). Pewnej, niewielkiej czêœci py³ku nie uda³o siê oznaczyæ ze wzglêdu na jego znaczny stopieñ zniszczenia w procesie trawienia. Na podstawie wyników analiz palinologicznych sporz¹dzono, dla ka dego roku badañ, taœmê pokarmow¹ dla Osmia rufa (Tab. 1, 2). S³owa kluczowe: murarka ogrodowa, Osmia rufa, roœliny pokarmowe, analiza py³kowa odchodów, trawienie py³ku.