Studies of honey bee species in lucerne seed production (Medicago sativa L.)

Introduction

Lucerne (Medicago sativa L.) called, 'queen of forage crops', is one of the oldest among cultivated fodder crops also known as alfalfa means best fodder as lucerne is most important cosmopolitan forage crop. Lucerne belongs to family leguminosae and genus Medicago. This genus includes 65 species native to Europe, Asia and Africa. Lucerne is propagated or reproduced by seed. Once established, lucerne gives fodder for 3-4 years. Lucerne is a wonderful winter fodder crop and mainly grown where berseem fails. It is valued for its drought tolerance, high yield, protein content and digestible fibre and as such makes good forage for ruminants. It is used for grazing, hay, silage making, green manure and as cover crop (Burezq, 2021) [14] . It is valued for the ability to fix nitrogen during drought when other legumes are not nodulated or not actively fixing nitrogen (Johnson and Rumbaugh, 1981) [30] . Lucerne fixes 83 to 594 kg atmospheric nitrogen per hectare (Aganga, 2003) [1] . Its presence in rotation maintains the nitrates and organic matter in soil. Lucerne is also recommended for seeding mined soil to reduce erosion and increase forage value (Cogliastro et al., 1990) [21] . Lucerne contains approximately 5 times as much protein as sorghum and ample quantity of vitamin 'A'. This forage legume is palatable with 72 per cent dry matter digestibility. It is high in mineral content and contains at least 10 different vitamins such as pantothenic acid, riboflavin, biotin, folic acid, choline, pyrodoxine, vitamin B12, K and C. Lucerne contains 20.2 per cent crude protein, 16.2 per cent digestible crude protein, 30.1 per cent crude fibre, 1240 g calcium and 350 g phosphorous per 100 kg lucerne and metabolic energy 2.17 M cal per kg (Banerjee, 1978) [5] In India, area, seed and green forage production of lucerne is nearly 10 lakh hectares, 150 kg ha -1 and 65-130 tones per hectare, respectively. Scarcity of green fodder is one of the most important limiting factors for achieving "White revolution". The present feed and fodder resources of the country can meet only 48 per cent of the requirement with a vast deficit of 62 and 22 percent of green and dry fodder, respectively. Lucerne being a perennial fodder crop could play a major role in this direction. Insect pollination is essential for the sustainability of both agricultural and natural ecosystems since 87 per cent of flowering plants and nearly 35 per cent of crops worldwide are dependent on insect and other pollinators. Honey bee pollination enhances crop yield, seed production, and fruit quality, making them an eco-friendly and cost-effective means of improving cross-pollinated crops (Free, 1970) [23] . Alfalfa is considered an insectary crop because it attracts a large number of insect pollinators (Kulkarni and Kumar, 2020) [32] .

Floral biology of lucerne -The plant bears irregular bisexual flowers, usually purple, blue, violet, yellow, or white, about 1/4" to 1/3" long arranged in axillary racemes in an acropetal pattern. Each raceme contains 5-40 insectpollinated flowers, and the fruits are spirally twisted pods. The flower has one standard petal, two wing petals, and two fused petals forming the keel. The flower contains ten anthers in diadelphous condition (Brunet et al., 2019) [12] . Alfalfa flowering extends over a relatively long period, with all flowers in a bloom opening within about one week (McGregor, 1976) [35] . The unpollinated flower remains open for seven days, but very soon after pollination it wilts and falls off.

Pollination & Fertilization of alfalfa -Standard petal of floret acts as a platform for bees to land on it. The lucerne flower is so constructed that the style and stamens are enclosed in a tense position within the keel. When this tension is released the style and stamens spring forward, causing the pollen (sexual column) to be distributed and the stigma to strike with some force against the standard petal. This process, referred to as "tripping" the flower, allows the pollen to become available for cross and self-pollination. Alfalfa pollen is moist, sticky, heavy and coalescent into large lumps and cannot be transferred by wind, only by pollinating insects. There is self-pollination in alfalfa, however, as stated by Tysdal (1946) [49] , in field conditions less than one percent of flowers pollinated in such a way produce seed. The available bee forage resources in the lucerne flower are pollen and nectar. When a honey bee enters its proboscis into a flower and triggers its opening mechanism (tripping), the stamens strike the bee's head, depositing pollen, while the stigma receives pollen from another flower, enabling cross-pollination. Thus the bee both trips and cross-pollinates the flower (Butler et al., 1956) [15] . opening of the flower is rapid and frequent, stylus strikes the honey bee and throws it away from flower or its proboscis gets stuck between anthers and the honey bee must spend a lot of effort to free its head out of the flower. As this mechanism can be forceful and inconvenient, nectar-collecting honey bees often avoid tripping the flowers and instead extract nectar by squeezing their proboscis between petals without tripping the flowers. As a result, they obtain nectar without pollinating the flowers.

Alfalfa potential for seed production -Although alfalfa produces an enormous number of flowers (about 3.3 billion flowers ha⁻¹) due to its genetics, only a fraction develop into seed-bearing pods (Tysdal, 1946) [49] . Based on estimates of pod set and seed production, potential seed yields of up to 5,000 kg ha⁻¹ have been suggested (Lesins, 1950) [34] , while pod formation in 46.7% of flowers could result in about 2,000 kg ha⁻¹ of seed yield (Pedersen et al., 1956) [37] . However, such yields are rarely achieved under field conditions primarily due to poor pollination of alfalfa flowers which is a result of morphological traits of flower, low self fertilization and insufficient presence of adequate pollinators. Practical seed yields are reported to be 20-30 times lower than the theoretical potential (Stjepanović, 1981), with average yields ranging from 250-300 kg ha⁻¹ and maximum yields reaching 800-1000 kg ha⁻¹ under favorable agro-ecological conditions (Erić, 1995). Pollinators have a vital role in pollinating the lucerne crop. Cross-pollination in lucerne contributes to 89 per cent seed yield (Somerville, 2002) [45] . Reproductive success in lucerne is influenced significantly by pollinator abundance, their foraging behaviour and pollination characteristics (through the provision of pollen and nectar). Alfalfa, is cross-pollinated because of self-incompatibility and selfsterility. One of the major problems in alfalfa seed production is Insufficient pollination and variation in seed yield, which is heavily affected by environmental conditions. The years with higher rainfall are more favourable to solitary bees, while the dry years are ideal for honey bees. Pollination is mainly dependent on honey bees and other solitary bees. By keeping all this in a view viz. economic and ecological importance of lucerne, problems associated with pollination and its yield potential, investigation was undertaken to study diversity of flowers visitors and their abundance in lucerne seed production plot and impact of managed bee pollination on seed yield obtained by use of honey bee colonies.

Materials and methodology 2.1 Materials

• Seed -The seeds of lucerne RL-88 variety were used for studies on pollinators in seed production • Honey bees-Honey bee hive of Apis mellifera, Apis cerena indica and Trigona irridipenis in lucerene crop will be placed inside the cage house • Pollination cages -Nylon nets will be used to make pollination cages. The cages of nylon mosquito nets(1 mm) size will be erected over the crop at the commencement of flowering.o erect pollination cages

Methodology 2.4.1 Species diversity of flowers visitors

A total of 1000 m 2 area of lucerne field was selected and the observations pertaining to insect flower visitors and their diversity on lucerne flowers recorded once a week starting from 11 th standard meteorological week SMW to 18 th standard meteorological week SMW during summer 2025 using transect sampling method. The observations were recorded at three hourly intervals (9.00-12.00 hrs, 12.00-15.00 hrs and 15.00-17.00 hrs) on a selected day of an individual week. The traversing distance was up to 10 m in one direction and total distance of 30 m (3 transect lines) was covered in a span of five min. A total of 24 such transect walks (10 meters each) were made throughout the flowering period and all the flower visiting insects were counted and recorded in a field register. Representative specimens of each insect visitor were caught with a hand net, pinned, labelled and repository of lucerne flower visiting insects was prepared which is later identified with the help of specialist.

To study effect of managed bee pollination seed yield of lucerne

The cages of the size 10 m × 10 m were made by using mosquito net of 288 mesh with four replications for each treatment. After crop maturity and seed setting seed yield, no. of pods per receme, test weight and germination test are recorded

Seed yield (kg/ha) -Inflorescences from all five replications of each treatment were harvested separately. Seeds were threshed, cleaned, and weighed for each replication. The average seed weight was expressed as kg/plot and later converted to kg/hectare Number of pods per raceme (P/R)-Five inflorescences were randomly collected from all four replications of each treatment. Pods were separated and counted individually, and the average value was expressed as number of pods per raceme (P/R).

Weight of 1000 seeds (Test weight)

Germination test -Germination tests for each treatment were conducted in four replications of 100 seeds following ISTA procedures (Anon., 2014) [4] using the above paper method. Final germination count was recorded on the fourth day based on normal seedling emergence and growth, and results were expressed as percentage seed germination.

ICBR ratio -The incremental cost-benefit ratio (ICBR) refers to a financial metric that evaluates the additional costs incurred in treatment (Appendix II) compared to the additional benefits gained (Vladica, 2018) [52] .

B:C ratio

The Benefit-Cost ratio (B:C ratio) is a financial metric used to access the profitability of a treatment. It compares the total present value of benefits generated by treatment to the total present value of cost. The B:C ratio of the respective treatment was calculated by using the following formula, (Akila and Babu, 1994) [3] 3. Results and discussion

Studies on species diversity of flowers visitors in lucerne seed production

Studies on the species diversity of flower visitors in lucerne fields are essential for improving crop production, supporting biodiversity conservation and promoting sustainable agriculture. The flower visitors of lucerne composed of 905 individuals of 27 different species which belonged to 22 genera, 12 families and 3 orders. These insect visitors visited the lucerne flowers for both pollen and nectar and did not cause any harmful damage to flowers of lucerne plant. The flower visitors predominantly belonged to the order hymenoptera (79.52%) followed by lepidoptera (11.09%), diptera (9.39%). The species were continuously active and spent most of the time of the day in visiting lucerne flowers and found as effective pollinators. Among 27 flowers visitors, 13 species of 4 families belongs to hymenoptera order. Honey bees included A. dorsata, A. cerana and A. florea, A. mellifera. Among honey bee species dominant species was A. florea (40 The results of present investigations are in line with Chavhan and Shashidhar (2015) [19] , Kulkarni and Kumar (2020) [32] , Rauf et al. (2021) [41] , Kumar et al. (2024) [33] and, who found that the majority of floral visitors belonged to Hymenoptera, followed by Lepidoptera and Diptera. The dominance of honey bees in the present study is consistent with the reports of Vansell and Todd (1946) [51] , Bieberdorf (1949) [7] and Ciurdarescu (1972), Cecen (2008) [17,18] and Jones and Forster (2012) [31] , who concluded that honey bees were the most frequent visitors to alfalfa flowers. Under Indian conditions, Gachchinamath (1983) [24] , also recorded A. cerana and A. florea as predominant floral visitors in lucerne forming more than half of the bee population. Studies conducted in Central Serbia for ten long year (1997-2007) it was found that honey bees were dominant compared to all other pollinators. In contrast, Hadfield and Calder (1936) [26] , Gurr (1955) [25] and Wojtowski (1965) [53] reported that Bombus terrestris was a more effective pollinator of lucerne than the honey bee. Brunet and Stewart (2010) [13] found that three species of the genus Bombus visited alfalfa flowers significantly more frequent and tripped them much better than the alfalfa leafcutter and honey bees. Conversely, Jevtic and Radovic (2001) [29] reported that honey bees formed only 1.1 per cent of pollinators. The presence of syrphid flies (7.82%) as significant floral visitors corroborates the findings of Kulkarni and Kumar (2020) [32] and Chavhan and Shashidhar (2015) [19] , who recorded syrphids as an important component of lucerne pollinator communities in India.

The role of non-Apis bees, syrphid flies and butterflies, though numerically less abundant, cannot be overlooked due to their documented efficiency in flower tripping and potential to enhance seed set. Studies by Vansell and Todd (1946) [51] Stephen (1960), Bohart (1962) [9] , Olmstead and Wooten (1987) [36] and Cecen (2008) [17,18] identified leafcutting bees (Megachile rotundata) and alkali bees (Nomia melanderi) as efficient pollinators. Rauf et al.

(2021) [41] emphasized the high pollination efficiency of solitary bees such as Megachile cephalotes and Amegilla spp. Bohart and Kmowlton (1960) [8] showed that these two species can be manipulated, i.e. they can be transported and domesticated in the alfalfa fields. As claimed by Mc Gregor (1976) [35] alkali bees and alfalfa leafcutters are more efficient pollinators than honey bees as their primary motive for visiting alfalfa flower is collecting pollen used to feed the brood. Bosch and Kemp (2005) [10] determined that alfalfa leafcutter can increase seed yield if brought organized to the seed crop field. Ciurdaresku (1974) [20] claimed that honey bee trips from 3.4 to 9.5% of visited flowers, which is much less efficient than some solitary bees which trip 45-100% of visited flowers. Tucker (1956) [48] pointed that honey bees which forage nectar trip only 0.3% of visited flowers. He gives great importance to young nectar foragers, citing the fact that they trip 70-85% of visited flowers during their "learning process". These findings emphasizing that lucerne pollination benefits from a diverse pollinator community rather than reliance on a single bee species.

Impact of managed bee pollination on seed yield parameters of lucerne

Managed bee pollination, specifically using honeybees, can have a significant positive impact on both the quantity and quality of lucerne seeds. Precise and efficient pollination process by managed honey bees tends to enhance seed uniformity and quality, leading to a higher percentage of viable and marketable crop seeds, ultimately benefiting the seed production industry. The data pertaining to various qualitative and quantitative parameters viz. lucerne seed yield (Kg/ha), Number of pods/ racemes (P/R), 1000 seed weight (g), Germination (%) are studied. The results on the effect of different pollination treatments on seed yield and yield attributes of lucerne revealed significant differences (Table 3). Among the treatments, open pollination (T4) recorded the highest seed yield (351.23 kg/ha), number of pods per raceme (19.1), 1000seed weight (3.76 g) and germination (95%). This was followed by A. cerana pollination (T1), which produced 297.6 kg/ha seed yield, 16.45 pods per raceme, 3.58 g test weight and 93.5% germination. T. iridipennis pollination (T3) also showed moderate improvement over the control with 254.82 kg/ha seed yield, 11. findings that insect pollinators are crucial for successful pod set and higher seed yields in lucerne. Gregor Similarly, Bieberdorf (1949) [7] and Rincker et al. (1988) [43] reported that tripping and insect-mediated cross-pollination are prerequisites for seed set in lucerne, which is consistent with the present findings. Jevtic and Radovic (2001) [29] reported that in the plot without honey bees (Apis mellifera carnica) 23.3 kg ha -1 and in cages with bees 158.9-198 kg ha -1 and in open pollination 304 kg ha -1 of seed yield were obtained. Cecen et al. (2008) [17,18] , using isolated cages, found that the bumblebee (Bombus terrestris L.) is more successful pollinator of alfalfa compared to the honey bee. In cages with bumblebees, the average seed yield was 56.48 kg ha-1, while in cages with honey bees, 49.2 kg ha-1 and with free pollination 66.19 kg ha-1 was obtained.. The higher seed yield under open pollination and A. cerana also confirms the reports of Vansell and todd (1946) [51] and Olmstead and Wooten (1987) [36] , who documented that seed yields were greatly enhanced with honey bee activity and

drastically reduced in pollination-excluded plots.

Comparable observations were also made by Gachchinamath (1983) [24] and Hosamani et al. (2020) [28] , who highlighted the role of A. cerana in significantly increasing lucerne and onion seed yield, seed weight and germination. The present study further aligns with Pushpalata (2024), who found that A. cerana pollinated plots outperformed A. mellifera and T. iridipennis in terms of yield and seed quality parameters in onion. Findings of Singh and Dharmwal (1970), Benedek and Gall (1972) [6] and Pudasaini et al. (2014) [39] , demonstrates that insect pollination not only increases seed yield but also improves seed viability and vigor. Likewise, Pashte et al. (2015) emphasized that A. cerana pollination enhances seed quality traits such as test weight and germination percentage in sesamum and other crops, which parallels the present findings. These findings reinforce earlier reports (Cane, 2002; Divija and Jayanthi, 2022) [16,22] that lucerne is a strictly cross-pollinated crop requiring active bee visitation and that bee pollination enhances both quantitative (yield) and qualitative (seed quality, germination) aspects of production. Previously, Kulkarni and Kumar (2021) [32] reported that open pollination along with solitary bees produced significantly more seeds as compared to honeybees and pollination-exclusion flowers. Similarly, Breazeale et al. (2008) [11] , Brunet et al (2019) [12] and Pitts-Singer et al. (2011) [38] concluded worldwide that higher abundance of Megachile bees has been linked with enhanced alfalfa seed production due to their superior pollination efficiency. Many authors claim that the honey bee is not very successful in the pollination of alfalfa (Benedek, 1976; Holm, 1984) [6] , primarily due to the anatomy of the flower. In addition, it was found that many other bee species are much more efficient pollinators of this important forage crops (Richards, 1984; Cane, 2002, Stanisavljević et al., 2003, etc.) [16,42,46] . However, those bee species have their specific drawbacks as they are highly dependent on meteorological conditions and anthropogenic factors. In addition, alfalfa is grown on large areas where there are no natural habitats for spontaneous pollinators. From the above stated, it can be concluded that the pollinating of alfalfa cannot be left to only one species, i.e. spontaneous pollinators, because of their low abundance and frequency and high dependence on various factors.

Conclusion

The studies on species diversity indicated that pollinator faunas of lucerne composed of 905 individuals of 27 different species which belonged to 22 genera, 12 families and 3 orders. The flower visitors belonged to the order hymenoptera (79.52%), lepidoptera (11.09%) and diptera (9.39%). Among five pollination treatment, best treatment was open pollination that gave highest seed yield (351.23 kg/ ha) with highest germination percentage and seed weight which was at par with A. cerena followed by pollination with T. iridipennis. A. mellifera yielded low seed with less germination percent than other honey bee pollination. However no pollination gave negligible seed yield indicating self-pollination in lucerne is not economically beneficial. Hence this study revealed Alfalfa, as practically only xenogame species in which pollinators play an important role in increasing the yield of lucerne. The honey bee is not as effective in pollination of alfalfa as are some solitary bees, but its advantage is in the number of colonies and the ability to manipulate them easily. In the pollination of alfalfa significantly greater importance is of pollen forager bees compared to the nectar foragers. Results of open pollinated field suggests adequate pollination by diversity of flowers visitors viz. honey bees, wild bees, syrphid flies, butterflies is needed for a significant increase in the seed yield with enhanced germination.

References

1. Aganga G. Lucerne, Lablab and Leucaena leucocephala Forages: Production and Utilization for Livestock Production. Pak. J. Nutrition. 2003;2(2):4653.
2. Aguiar WM, Melo GAR, Gaglianone MC. Does Forest phisiognomy affect the structure of orchid bee (Hymenoptera, Apidae, Euglossini) communities? A study in the Atlantic forest of Rio de Janeiro state, Brazil. Sociobiology. 2014;61:68-77.
3. Akila S, Babu PCS. Release of different dose of Trichogramma and its effect on internode borer, yield and quality of sugarcane. Sugarcane (London). 1994;2:22-23.
4. Anonymous. International rules for seed testing by international seed testing association (ISTA). Seed Science and Technology. 2014;27:215.
5. Banerjee GC. Principles of animal nutrition and feeding practices. Chpt., Animal Feed Stuffs. 1978:1-20.
6. Benedek P, Gall E. The effect of insect pollination on seed onion with observation in the behaviour of honey bees crop. J. Apic. Res. 1972;11:175-180.
7. Bieberdorf GA. Some observations on pollination of alfalfa hay. Academy of science for proceeding of the Oklahoma. 1949:49-51.
8. Bohart GE, Knowlton GF. Managing alkali bees for alfalfa pollination. Logan, Utah State Univ. Extension Services Leaflet. 1960;78:1-8.
9. Bohart GE. Management of wild bees for the pollination of crops. Annu. Rev. Entomol. 1962;17:287312.
10. Bosch J, Kemp WP. Alfalfa leafcutting bee population dynamics, flower availability and pollination rates in two Oregon alfalfa fields. Journal of Economic Entomology. 2005;98(4):1077-1086.
11. Breazeale D, Fernandez G, Narayanan R. Modeling pollination factors that influence alfalfa seed yield in north-central Nevada. J. Cent. Eur. Agric. 2008;9:107116.
12. Brunet J, Zhao Y, Clayton MK. Linking the foraging behaviour of three bee species to pollen dispersal and gene flow. Plos One. 2019;14:e0212561.
13. Brunet J, Stewart CM. Impact of Bee Species and Plant Density on Alfalfa Pollination and Potential for Gene Flow. Hindawi Publishing Corporation, Psyche. 2011;Volume 2010(Article ID 201858):7 pages.
14. Burezq H. Reproductive ecology of forage alfalfa (Medicago sativa L.): Recent advances. Plant Reprod. Ecol. 2021;579:100-640.
15. Butler George D, Todd Frank E, Tuttle Donald M. Alfalfa pollination in Arizona. A. Progress Report. Agr. Expt. Sta., University of Arizona (Tucson, AZ). 1956. International Journal of Advanced Biochemistry Research https://www.biochemjournal.com
16. Cane JH. Pollinating bees (Hymenoptera: Apiformes) of U.S. alfalfa compared for rates of pod and seed set. Journal of Economic Entomology. 2002;1:22-27.
17. Cecen S, Gurel F, Karaca A. Impact of honeybee and bumblebee pollination on alfalfa seed yield. Acta Agriculturae Scandinavica, Section B -Soil & Plant Science. 2008;58(1):77-81.
18. Cecen S. Impact of honeybee and bumblebee pollination on alfalfa seed yield. Acta Agriculturae Scandinavica, Section B -Soil & Plant Science. 2008;58(1):77-81.
19. Chavhan VT, Shashidhar V. Pollinator fauna of lucerne and their relative abundance in lucerne at Dharwad, Karnataka. Karnataka Journal of Agricultural Sciences. 2015;28(1):112-113.
20. Ciurdaresku G. Polination as an important factor for the yield alfalfa crops in Romania. Organizing committee XII international grassland congress. Sectional Papersplant introduction, Breeding and seed production - Moscow. 1974:60-69.
21. Cogliastro Alain, Gagnon Daniel, Coderre Daniel, Bhereur Pierre. Response of seven hardwood tree species to herbicide, rototilling and legume cover at two southern Quebec plantation sites. Canadian Journal of Forestry. 1990;20:1172-1182.
22. Divija SD, Jayanthi KPD. Pollination efficiency and foraging behaviour of honey bees and flies to onion Allium cepa L. Journal of Apicultural Research. 2022;61(5):688-694.
23. Free JB. Insect Pollination of Crops. 2nd ed. London Academic Press; 1970:544.
24. Uni. Agri. Sci., Dharwad (India); 1983.
25. Gurr L. A note on the efficiency of Bombus terrestris (L.) as a pollinator of lucerne (Medicago sativa L.). N.Z. J. Sci. Tech. 1955;37A:300.
26. Hadfield JW, Calder RA. Lucerne (Medicago sativa) investigations relative to pollination and seed production in New Zealand. N.Z. J. Sci. Tech. 1936;17:577-594.
27. Holm SN. Introduction and propagation of the leafcutter bee (M. rotundata) in Denmark. Proc. int. Sym. on Poll., 27-30 IX 1983. Versailles, Tasei, Jnied. 1984:455-460.
28. Hosamani V, Venkateshalu, Jagadeesha N, Reddy MS, Gangadarappa PM, Ravikumar B. Effect of Bee Pollination and Attractants on Quantitative and Qualitative Parameters of Onion (Allium cepa) Seed Yield and Quality. Int. J. Curr. Microbiol. App. Sci. 2020;9(2):1885-1892.
29. Jevtic G, Radovic J. The importance of honey bee (Apis mellifera carnica) in pollination and seed yield of alfalfa (Medicago sativa L.). Ministry of Education, Science and Technology Development of the Republic of Serbia. 2001:11-24.
30. Johnson DA, Rumbaugh MD. Nodulation and acetylene reduction by certain rangeland legume species under field conditions. J. Rang. Mgmt. 1981;34(3):178-181.
31. Jones P, Forster W. Observations on the pollination of lucerne (Medicago sativa Linn.) N. Z.J. Agric. Res. 2012;8(2):340-349.
32. Kulkarni Narendra S, Kumar Vinod. Studies on Insect Fauna of Lucerne (Medicago sativa) and Their Impact on Seed and Forage Yield in Lucerne Medicago sativa. IGC Proceedings (1989-2023). 2020. Available from: https://uknowledge.uky.edu/igc/23/2-2-1/
33. Kumar S, Keerthi MC, Mahesha HS, Arunkumara CG, Shivakumara KT, Saini RP, Yadav VK. Foraging behaviour and pollinator dynamics in lucerne crop of the Bundelkhand region: Implications for crop management in the subtropics. Animal Biology. 2024;74(4):401-413.
34. Lesins K. Investigations into seed setting of lucerne at ultuna Sweden. Ann. Roy. Agr. Col. (Uppsala: Sweden). 1950;17:441-483.
35. Mc-Gregor SE. Alfalfa in: Insect pollination of cultivated crop plans. Agricultural Handbook, US Department of Agriculture No. 496. 1976:64-75.
36. Olmstead AL, Wooten DB. Bee pollination and productivity growth: The case of alfalfa. American Journal of Agricultural Economics. 1987;69:56-63.
37. Pedersen MW, Petersen HL, Bohart GE, Levin MD. A comparison of the effect of complete and partial crosspollination of alfalfa on pod set, seeds per pod and seed weight. Agron. Jour. 1956;48:177-180.
38. Pitts-Singer TL, Cane JH. Bee pollination of lucerne: problems and solutions. 2011.
39. Pudasaini RB, Thapa VR, Poudel PR. Effect of Pollination on qualitative characteristics of rapeseed in Chitwan, Nepal. International Journal of Biological, Biomolecular, Agricainursl. Food and Biotechnological Engineering. 2014;8(12):65-68.
40. Univ. Agric. Sci., Rahuri (India); 2024.
41. Rauf A, Saeed S, Ali M, Nadeem Tahir MH. Comparative efficiency of native insect pollinators in reproductive performance of Medicago sativa L. in Pakistan. Insects. 2021;12(11):1029.
42. Richards KW. Alfalfa leafcutter bee management in Western Canada. Publication 1495/E Agricultural Canada: Ottawa, Canada. 1984:53.
43. Rincker CM. Alfalfa seed production in the Pacific Northwest. In: Proceedings Annual Farm Seed Conference. Am. Seed Trade Association, Kansas City, Mo. 25 November. 1988:13-19.
44. Singh JP, Dharamwal SS. The role of honey bees in seed setting on onion at Pant Nagar, Dist. Nainital, Uttar Pradesh, India. Indian Bee Journal. 1970;32(1/2):23-27.
45. Somerville D. Lucerne pollination (Agfact DAI-134). New South Wales Department of Primary Industries. 2002.
46. Stanisavljević Lj, Krunić M, Brajković M, Jevtić G. Značaj medonosne i drugih vrsta pčela za oprašivanje gajenih biljaka. XI Naučno savetovanje sa međunarodnim učešćem, 6-9. februar, Poljoprivredni fakultet, Zemun, Zbornik radova. 2003:82-88.
47. Tucker KW. Honey bee pollinators of alfalfa in Minnesota. 15th Alfalfa Imp. Conf. Rpt. St. Paul, Minn. 1956:32-33.
48. Umaerus M, Akerberg E. Pollination and seed setting in red clover and lucerne under Scandinavian conditions. Herb. Abstr. 1959;29:157-164.
49. Vansell GH, Todd FE. Alfalfa tripping by insects. Amer. Soc Agron. Jour. 1946;38:470-478.
50. Vladica V. How to calculate incremental cost benefit ratio? ResearchGate. 2018. Available from: https://www.researchgate.net/post/How-to-calculateincremental-costbenefitratio/5bb2b2bfeb038993023a07db/citation/download
51. Wojtowski. Zastosowanie blonkowek pszolowatych rodzaju Bombus latr. (Hymenoptera: Apoidea) do zapylania planteji nasiennych rostin motylkowych. Roozn. Wyz. Szk. Roln. Poznen. 1965;24:223-274.