Concentration of Heavy Metals in Pollen and Bees Osmia bicornis L. in Three Different Habitats in the Łowicz District in Central Poland

Abstract

The aim of our research was to compare the levels of pollution in different habitats based on the concentrations of heavy metals found in pollen and the organisms of Osmia bicornis L. bees in three habitats: orchards, berry plantations, and urban habitats (near by power plant, landfill, residential areas, and heavy vehicle traffic). The concentration of Ag, Cd, Cu, Fe, Mn, Ni, Pb and Zn were determined. Samples were quantified using flame atomic absorption spectrophotometry (AAS), with evaluations being carried out three times for each sample. Bee nests were also analyzed concerning reproduction and the presence of parasites. There were no significant differences in the content of heavy metals Ag, Cd, Cu, Fe, Mn, Ni, Pb and Zn in pollen samples between the three habitat types. Bee samples differed only in their Zn content, which was significantly higher in orchards (2.67 mg/kg) than urban habitats (0.80 mg/kg) and berry plantation (0.94 mg/kg). Habitat type had no effect on most bee reproductive parameters (percentage of occupied nest tubes, population growth). Our results show that bees pollinating crops in, for example orchards or berry plantations are exposed to heavy metals to a similar extent to those urban areas.

1. Introduction

Studies on bioindication tend to focus on metal pollution and other chemical pollution [1]. Anthropogenic sources of heavy metals include urbanization and industry [2,3,4], road transport and dust from road traffic [5,6] and agriculture [7,8,9,10,11]. Various organisms can be bioindicators of pollution (for example, heavy metals) in a given region.

Heavy metals are released into the air in the form of dust and ash from many sources, they pollute both soil and water [11,12]. Metal contamination of the soil has a negative impact on the populations of earthworms and macrofauna, for example, ants [13] and beetles [14]. Contaminants accumulate on the surface of plants as a result of precipitation or sedimentation [15]. Pollutants are also absorbed and collected by plants and become of the pollen and nectar of these plants [16]. Pollinating insects eat contaminated food, which is how pollutants enter their organisms. Pollutants may have a negative impact on various groups of pollinators, for example, on butterflies [17,18], bumblebees [19,20], and solitary bees [2,5,21]. The level of heavy-metal pollution according to Xiaoyu Shi et al., (2023) is negatively correlated with the diversity and species richness of wild bees [22].

Wild bees are a major group of pollinators in the temperate zone [23]. The decline in the population of wild bees negatively affects the pollination of plants, and thus the production of fruits and plant seeds [24,25,26]. There is evidence that heavy-metal pollution is causing wild bee habitats to become fragmented or lost [19,21]. Moroń et al. [2] showed that with increasing heavy-metal concentration, there is a steady decrease in the number, diversity, and abundance of solitary, wild bees. At sites with high concentrations of heavy-metal contamination (the concentration of cadmium, lead and zinc in pollen collected by bees ranged from 6.7 to 9.3, 277.0 to 356.2, 440.1 to 592·4 (mg·kg⁻¹), respectively), a higher share of dead individuals of Megachile lignisea L. was observed [2], while for Osmia bicornis L. there was a steady decrease in the number of brood cells constructed by females and an increase in the proportion of dead [2]. Increasing environmental pollution with cadmium, lead and zinc negatively affects the wings size of red mason bee females [27]. Preliminary research has shown that they can also cause asymmetry in the bees’ wings [27].

Among the species of wild bees, common in Europe, bred and used to pollinate crops, is red mason bee Osmia bicornis (Linnaeus, 1758). syn. rufa (Hymenoptera; Megachilidae) [28,29,30,31,32,33,34]. For the purpose of analysis, O. bicornis can provide bees (body), pollen, and also soil, which they use to build chambers in tubes [35]. O. bocornis’ main advantage for environmental studies is their short flight range around the nest, approximately 600 m [36]. In addition, the diversity of provisions within the nests of O. bicornis is low [36], which indicates that it mainly collects pollen from plants available close to the nest. However, only a few studies have indicated the usefulness of the nest bundles of solitary bees for characterizing threats to insects inhabiting concentrated agricultural or urbanized areas. Peterson et al. [35] showed that, in an agriculturally intensive landscape, all the nesting and brood matrices in reeds (mud, leaves, brood, and pollen) contained different agrochemicals. Bees and wasps in agricultural areas are exposed to agrochemicals at all stages of their lives, at often high frequencies and in potentially lethal concentrations and can influence the higher mortality of larvae.

Heavy-metal contamination mainly affects urbanized and industrial areas. The level of heavy-metal contamination of crops in agricultural areas and the impact of pollution on solitary bee reproduction are still unknown. The aim of our research was to compare the level of pollution in different habitats based on the concentrations of heavy metals in pollen and the O. bicornis organism. The study assessed the concentrations of the heavy metals Ag, Cd, Cu, Fe, Mn, Ni, Pb and Zn in pollen collected from O. bicornis nests and bee bodies for three types of habitats: orchard, berry plantation, and urban areas, and presented the impact of the habitat on the population development of bees.

Although the aim of the study was not to compare O. bicornis bee and the honey bees as a bioindicators, this investigation may influence the spread of the use of O. bicornis in biomonitoring. Breeding O. bicornis bees is generally considered to be less laborious and less expensive than honeybees maintaining hives. The use of O. bicornis as a bioindicator may be particularly important in places that require frequent environmental monitoring.

2. Material and Methods

2.1. Study Area

The study was carried in 2017 out in the Łowicz district of Poland (Łowicz county 52°6′0″ N, 19°55′60″ E), which is a typical agricultural area. There is one municipal heating plant in the south of the region and a garbage dump to the west of the town of Łowicz. Boxes with O. bicornis nests and cocoons were placed at 27 different sites (Figure 1). One nest with 500 reed tubes and 500 O. bicornis cocoons was placed at each location. The nests were located in three types of habitats (Figure 1 and Supplementary Materials S1):

(1)

berry plantations (currants, Kamchatka berry, blueberry), 8 sites, with an area of 1.78–3.6 ha, in a plantation structure—row spacing of plants 1.5–3 m, distance between plants 1–1.2 m, tree height 1.8–2 m,

(2)

orchards (apple trees, pear trees, plums trees), 10 sites, with an area of 1.6–4.1 ha, in a orchard structure—row spacing of plants 3–4 m, distance between plants 2–3 m, plants height 3.5–4 m;

(3)

urban areas (9 sites located near a power plant, landfill, residential areas, and heavy vehicle traffic).

The minimum distance between the different site types (nests) was 500 m.

Figure 1. Distribution of the study sites in Łowicz district in central Poland.

Figure 1. Distribution of the study sites in Łowicz district in central Poland.

2.2. Nest Construction and Material

The boxes containing nests and bee cocoons were placed at the sites during the time when O. bicornis naturally emerged, i.e., in mid-April. The nests were placed with the opening facing south or south-west at a height of 1 m, in places with moderate sunlight.

The female-to-male ratio in the experimental population was reviewed, and the average was 1:1. We assumed that one female would use two tubes for their nest [28]. In each habitat were placed 500 cocoons and 500 reed tubes (5 packages of 100 tubes each, with a length of 20 cm and an inside diameter of 6–8 mm) in each habitat.

2.3. Collection and Conservation of Samples

Pollen: Pollen was collected once, three weeks after the O. bicornis started flying. Three to four nest tubes were colleted from each nest/site. There were about 10 chambers containing pollen for each nest tube. The pollen was extracted from the chambers, after removing bee larvae. The pollen from all the chambers form each nest/site was mixed and formed the aggregate sample. Then, 25 g of pollen was placed in plastic containers and stored at −25 °C before the analysis.

Bees: O. bicornis cocoons were collected from each nest/site. The bees were removed from the cocoons using a scalpel and allowed to excrete feces. Then, 20 bees (10 females and 10 males) were taken and placed in plastic containers and frozen at −25 °C.

2.4. Preparation of Samples and Heavy-Metal Analysis

Each sample was analyzed by three parallel measurements. In order to obtain precisely dried and homogenized samples, both pollen and bees were triturated and placed on RADWAG WPX 50S moisture balances, where they were dried to a constant weight. Each sample was weighed at 1 g (to the nearest 0.10 mg) and samples of biological material were weighed in Teflon dishes using a RADWAG WAS 220/X analytical balance. The samples were covered with 5 cm³ of spectrally pure 69% nitric acid (TRACEPUREMD Millipore Corporation Darmstadt, Germany). These sample preparations were mineralized in an ANTON PAAR MULTIWAVE 3000 microwave digestion unit for 30 min. The whole process lasted two hours. The samples were analyzed quantitatively using flame atomic absorption spectrophotometry (AAS). Elements that were analyzed included Ag (silver), Cu (copper), Mn (manganese), Fe (iron), Ni (nickel), Cd (cadmium), Pb (lead) and Zn (zinc).

The results of the analysis were verified using certified reference materials—IAEA-336, the International Atomic Energy Agency’s Analytical Quality Control Services, Austriaand CRM 482 Commission of the European Communities‘s Community Bureau of Reference (BCR). A Spectra AA-110/220 from Varian (Australia) was used for the analysis. The limit of detection (LOD) was 1 ppb [37,38,39,40].

2.5. Nest Analysis and Bees’ Reproductive Parameters

Nests were selected for analysis in the laboratory at the end of September. Each tube was cut open with a scalpel and the number of breeding cells in which there were cocoons, dead larvae, and pupae, or parasites was recorded. In this study, we determined the parameters of the population, which were used in previous studies [41]:

a. Percent of tubes occupied by bees in relation to all tubes in the nest.

b. Mean number of cocoons per nest tube was calculated according to the following formula.

$$Cff=\frac{Nc}{Nt}$$

(1)

C_ff—mean number of cocoons per nest tube;

N_c—total number of cocoons harvested;

N_t—number of nest tubes.

c. Mean number of dead larvae and pupae per nest tube was calculated according to the below formula.

$$L=\frac{Nl}{Nt}$$

(2)

L—mean number of dead larvae and pupae per tube nest;

N_l—total number of dead larvae and pupae in nest;

N_t—number of nest tubes.

d. Mean number of parasites and kleptoparasites per nest tube was calculated according to the following formula.

$$P=\frac{Np}{Nt}$$

(3)

P—mean number of parasites and kleptoparasites per nest tube;

N_p—total number of chambers occupied by parasites and kleptoparasites in nest;

N_t—number of nest tubes.

e. Population growth rate was calculated according to the below formula.

$$Pgr=\frac{Nf}{Ni}$$

(4)

Pgr—population growth rate;

Nf—final number of cocoons (cocoons harvested after the season—offspring generation);

Ni—initial number of cocoons (cocoons placed at the site in spring—parental generation).

2.6. Statistical Analysis

For statistical analysis, R 4.1.2 with RStudio (R Core Team 2021) was used. In all tests, the level of significance was α = 0.05. The normality of data distribution was tested using the the Shapiro–Wilk test. The Kruskal–Wallis test with Holm correction for multiple comparison was used to check the differences between groups (package ‘agricolae’). For the data preparation, the packages ‘dplyr’, ‘tidyr’ and ‘tibble’ were used; the data visualization was carried out using the ‘ggplot2′ package.

3. Results and Discussion

3.1. Heavy-Metal Level in Samples of Pollen and Osmia bicornis Body

All the analyzed material samples (pollen and bees) from the three types of habitats exhibited a similar content of the metals as shown in Figure 2 and Figure 3. Honey bees have been the subject of most of the research on the impact of heavy-metal impact on pollinating insects [4,9,14,42,43,44,45,46,47,48,49] and thus we compare our results primarily to the heavy-metal content found in honey bees and their products.

3.2. Pollen Samples

The average concentration of heavy metals in pollen samples from all habitats was as follows: Fe > Mn > Cd > Cu > Zn > Ni > Pb > Ag (Figure 2).

Ag concentration in pollen did not differ across the three types of habitats and averaged 0.26 mg/kg in berry plantations, 0.24 mg/kg in orchards, and 0.36 mg/kg in urban habitats.

Arslan and Arıkan [50] showed that there was no statistical difference between the averages of Zn, Fe, Cd, Cr, or Mn in pollen samples from honey bee hives located away from traffic at varying distances, while the average content of copper and lead turned out to be statistically significant. According to the same authors, the greatest accumulation of cadmium occurs on side of the motorways, which confirms that the main source Cd is roads in close proximity to hives.

This study showed that across the berry plantations, orchards, and urban habitats, the average pollen Cd level did not vary (0.06–0.07 mg/kg). Our research also found that the level of this element in bee pollen was several times higher than that described by other authors in other countries, for example, in Finland [45] or in Turkey [43]. Formicki et al. [51] for the Małopolskie Voivodship in southern Poland demonstrated that there was a large variation in the content of cadmium in pollen depending on the location of the apiary—from 0.026 mg/kg to 0.092 mg/kg. This was similar to the findings of Roman et al. [4] for an industrial area in Lower Silesia in south-western Poland (0.001–0.092 mg/kg). In our research, the lowest concentrations of cadmium were found in berry plantation (0.04 mg/kg) and the highest concentrations in the urban habitats (0.10 mg/kg).

We assume that the sources of such high cadmium concentrations in urban areas were primarily coal stoves, which are still popular in households, and road traffic. The highest level of Cd was found in pollen collected from the area of a housing estate (0.1 mg/kg). Cd concentrations in other urban locations were similar and ranged from 0.05 to 0.07 mg/kg. Plant protection products that contain cadmium as an active substance or as a component of formulations may be responsible for high levels of cadmium in cropping areas [52]. Despite a much higher concentration of cadmium in comparison to other tests, its level in pollen did not exceed the International Food Standard values in the Codex Alimentarius 1995, amended 2019 [53].

Pesticide formulations also contain Ni and Pb [13]. Formicki et al. [18] found Ni levels in pollen of between 3.61 mg/kg and 8.41 mg/kg. The lowest concentration of Ni was found in the berry plantations (0.27 mg/kg) while the highest content was, in the orchards (6.39 mg/kg, average for berry plantation—1.76 mg/kg; orchards 1.75 mg/kg; urban 1.81 mg/kg). Much lower Ni content was found by Bayir and Aygun [43] in urban habitats in Turkey, where the maximum concentration amounted to only 0.384 mg/kg.

The Pb content did not differ significantly across the berry plantations, orchards, and urban habitats (0.42 mg/kg, 0.27 mg/kg and 0.35 mg/kg, respectively). These values were higher than those reported by other authors [3,43,45], and above concentrations recommended for most plant food in the International Food Standard values in Codex Alimentarius 1995, amended 2019 [53].

The Cu content also did not significantly differ across the three types of sites (10.97 mg/kg for berry plantations, 10.65 mg/kg for orchards and 9.94 mg/kg for urban habitats). We found both the lowest and the highest Cu concentrations in pollen from orchard areas 5.39 mg/kg and 22.17 mg/kg, respectively. This was much higher than the level of Cu found in urban habitats in Turkey (5.16–7.28 mg/kg, [43]) and somewhat similar to that recorded in Finland (7.90–9.90 mg/kg, [45]).

In our study, the lowest Fe values in pollen were found in urban habitats, which averaged 94.43 mg/kg. Higher values were found for orchards (303.18 mg/kg) and berry plantations (165.99 mg/kg). The pollen Fe contents in Turkey for urban habitats ranged from 73.18 mg/kg to 96.95 mg/kg [43].

We found no significant differences in the amounts of Mn in pollen for berry plantations, orchards, or urban habitats (84.70 mg/kg, 60.57 mg/kg, and 54.76 mg/kg, respectively), while the level of this element was several times higher than given by Bayir and Aygun for urban habitats in Turkey [43]. In contrast to the results obtained from Turkey [43], in our results the average Mn content of pollen in agricultural areas was higher than that from urban habitats. Fakhimzadeh and Lodenius [45] also found no significant statistical differences for the Mn values in pollen samples across industrial, urban or rural areas.

The Zn content was, again, broadly similar across the three types of sites (6.28 mg/kg in berry plantations, 5.65 mg/kg in orchards, and 5.62 mg/kg in urban habitats). The lowest and highest concentrations of Zn were found in orchards (4.29 mg/kg and 8.48 mg/kg, respectively). In Turkey, the Zn content in pollen samples ranged from 10.25 mg/kg to 20.27 mg/kg [43]. The average for our results for Zn were two to four times lower than those reported in Turkey (10.25 mg/kg to 20.27 mg/kg, [43]) and much lower than those found in different regions in Poland in 2013 (75.2–159.3 μg/g, [51]). We assume that this is due to differences in the studied areas and the potential emitters of heavy metals.

3.3. Bees Samples

The average concentration of heavy metals in the beessamples from all habitats was as follows: Fe > Cd > Cu > Mn > Zn > Ni > Ag > Pb (Figure 3).

The Ag concentration in the bees’ bodies did not differ across the three types of habitats, and averaged 0.36 mg/kg in berry plantations, 0.43 mg/kg in orchards, and 0.72 mg/kg in urban habitats.

Cd content in the bee bodies was similar for all three types of habitat. The average Cd values we found in the bee samples were 0.07 mg/kg for urban habitats, 0.08 mg/kg for orchards and 0.07 mg/kg for berry plantations. The lowest Cd value was found in urban habitats (0.068 mg/kg) and the highest in orchards (0.096 mg/kg). These values were within the reference values proposed by Guttiérrez et al. in 2015 (0.1 mg/kg) [54]. The Cd values in our results were close to those obtained by Roman in Poland in 2010 [55] for urban area (0.60 mg·kg⁻¹) and agroforestry areas (0.70 mg·kg⁻¹) in honey bee foragers, but much lower than they found in the agricultural areas in The Netherlands (0.23 mg/kg) [56]. Results from Turkey in 2022 [43] showed significantly higher concentrations of Cd in honeybee samples from urban areas (17.48–20.78 μg/kg) than in rural areas (9.52–13.25 μg/kg), although their values were several times lower than those in our study. Conti and Botrè, in 2001 [56] and Fakhimzadeh and Lodenius in 2000 [45], also reported lower Cd concentrations in bee samples from urban and industrial areas compared to samples from rural areas.

In our study, the Cu content in the bee samples was 14.81 mg/kg for urban habitats, 15.31 mg/kg for berry plantation and 15.31 mg/kg for orchards. These results were similar to those for urban areas in Turkey (13.43–17.18 mg/kg, [43]) and for the agricultural and urban area for the Netherlands (0.23 μg/g and 0.18 μg/g, respectively [57]). Our result showed lower Cu content than results from south-western Poland (22.00 mg/kg urban regions, 23.3 mg/kg agricultural–woodland regions [55]). DiFiore et al., in 2023 [58] showed higher Cu concentrations in agricultural areas in June–July (18–176 µg kg⁻¹), which may be the result of application for fungal pathogens control of crops. Silici et al., in 2016 [59] found that there was no effect of on the Cu levels in bees due to the proximity of a thermal power plant, but Taha et al., in 2017 [15], found that the Cu content in bees had higher values near a cement plant.

In our result, Fe content did not differ across the habitats, and had values of 71.26 mg/kg for berry plantations, 79.41 mg/kg for orchards, and 72.24 mg/kg for urban areas. The lowest Fe value was recorded at berry plantations, 64.77 mg/kg, while the highest was found in the orchards 92.59 mg/kg. Results from Turkey [43] showed higher Fe content in honeybee samples from urban areas (91.59 to 101.81 mg/kg) and from rural areas (82.46 to 96.35 mg/kg).

Our research found that Mn concentrations in the bodies of solitary bees did not differ across habitats, and ranged from 3.55 mg/kg in a blueberry plantations to 4.23 mg/kg for urban areas. The lowest Mn value was recorded for berry plantation, 2.95 mg/kg, and the highest in the orchards, 5.50 mg/kg. In 2016, Silici et al. [59] found no statistical differences between Mn values in honeybee samples collected at different distances from a CHP plant. Bayir and Aygun (2022), in Turkey, [43] revealed several times higher concentrations of Mn in honey bee samples from urban (max. 35.55 mg/kg) and rural areas (min. 15.63 mg/kg). Our results for the Mn content in bees were much lower for all habitats than those reported by Van der Steen et al., (2008) for the agricultural and urban areas in the Netherlands (162.4 mg/kg and 92.20 mg/kg, respectively) [57].

The mean values for Ni in the bee samples were quite similar across habitats, and ranged from 0.74 mg/kg for urban habitats to 0.86 mg/kg in berry plantations. The lowest and highest Ni values were both recorded for orchards, 0.16 mg/kg and 2.37 mg/kg, respectively. These values were almost twice as high as those obtained by Bayir and Aygun in Turkey in 2022 [43], in bees from urban areas, almost four times higher than in rural area. An Ni concentration above i above 0.3 mg/kg in bees is a disturbingly high concentration according to the proposed standards for honey bees by Gutiérrez et al. in 2013 [54]. Nickel has a wide distribution in the environment, being an essential component in almost 100 minerals. The industrial and commercial uses of nickel and nickel compounds are also numerous. Nickel is primarily used to manufacture stainless steel and nickel alloys, and has high corrosion and temperature resistance. Metals and alloys made from nickel are used widely in metallurgy, chemicals, and food processing. As one of the many trace metals widely distributed in the environment, nickel is released from both natural and anthropogenic sources. It can be found in air, water, soil, and biological materials [60]. Nickel is an essential microelement for plant growth [61]. As such, high Ni content as those found in our results, especially in orchards, are probably the result of the use of mineral fertilizers. Ni is also released by car brake abrasion, and vehicle corrosion (especially the car’s oil pump [62]) may be the reason for such high Ni levels in urban areas (roads, and cities).

The Pb content in the bee samples did not differ across habitats, and averaged 0.16 mg/kg for orchards, 0.11 mg/kg for berry plantations, and 0.10 mg/kg for urban habitats, which are all within the standard of 0.3 mg/kg defined as low pollution [54]. The lowest and highest Pb values were both recorded in orchards: 0.04 mg/kg and 0.45 mg/kg, respectively. In 2010 in Poland [55], and in 2022 in Turkey [43], the levels of Pb in honeybee samples were found to be higher in urban areas than agricultural–woodland and rural areas. In Italy, in 2023 [58], the Pb concentration in bees from urban, industrial, and agricultural habitats were 39, 49, and 29–40 μg·kg⁻¹, respectively, which is also higher than those for our study. The Pb values obtained in our study were significantly lower than those reported in Poland, (1.91–1.98 mg/kg [55]), in Italy, (0.14–0.52 mg/kg [63]) and in Turkey, (0.192–0.358 mg/kg [43]).

The Zn content in the bee samples in our study were significantly higher for orchards (2.67 mg/kg) than for urban habitats (0.80 mg/kg) or berry plantations (0.94 mg/kg). The highest concentration of Zn was found in orchards, at 5.57 mg/kg, which is much lower than the results reported from Turkey (29.93 mg/kg–44.58 mg/kg, [3]), from the Netherlands (96.80–98.50 mg/kg, [56]), and from Poland (approx. 20–45 mg/kg, [64]).

3.4. Bee Population Grow Rate in Urban, Orchards and Berry-Plant Habitats

In urban habitats and berry plantations bees occupied a similar percentage of nesting tubes, over 50%, while, in orchards, they occupied more than 60% of nesting tubes (

Table 1. Reproduction parameters of Osmia bicornis in three habitats—berry plantation, orchard and urban areas.

	Habitat
	Orchards				Berry Plantations				Urban
	Mean	Median	Min–Max	SD	Mean	Median	Min-Max	SD	Mean	Median	Min–Max	SD
occupied tubes (%)	62.0	74.7	22.3–85.7	26.0	52.9	65.8	4.74–87.6	38.3	51.9	33.3	8.4–97.6	38.7
cocoons/tube	6.75	6.76	4.9–8.83	1.36	6.76	7.32	4.22–8.01	1.25	6.03	5.93	5.21–7.41	0.881
death larvae and pupa/tube	1.73	1.56 B *	1.19–3.1	0.585	2.27	2.26 A	1.71–2.98	0.436	2.0	2.05 AB	1.57–2.38	0.389
chambers occupied by parasites/tube	0.901	0.36	0.22–2.86	1.01	0.342	0.405	0–0.71	0.245	0.848	0.84	0.29–1.88	0.632
population growth rate	3.97	3.87	0.85–7.55	2.11	3.11	3.6	0.22–5.61	2.46	3.09	1.93	0.438–6.05	2.58

* A, B: Different letters in the same line indicate significant differences (p < 0.05), Kruskal–Wallis test with Holm correction for multiple comparison.

). These were low results compared to studies on the urbanization gradient, where the percentage of settlements in urban and agricultural habitats was over 90% [41].

The percentage of occupied nests influenced the population growth, which was much higher in orchards (3.97) than in berry plantations or urban habitats (3.60 and 3.09, respectively). Orchards achieved a population increase of 3.17 [65], a result similar to ours for the same type of habitat. In urbanized habitats, the population growth rate of O. bicornis was 5.65 [41]. The amount and availability of pollen for O. bicornis in a mixed environment is greater than in monocultures, which ensures an abundance of pollen over a short time. This may have resulted in higher population growth in urban habitats than in orchards and berry plantations (Table 1).

Pollen collected by O. bicornis females showed no differences in heavy-metal concentrations, but we did notice some differences in larval mortality between habitats. Larval mortality in nests located in orchards was significantly lower (1.73) than in nests located in berry plantations (2.27). There was no difference in larval mortality between orchards (1.73) and urban habitats (2.0) (Table 1).

Moroń et al. in 2014 [21] found that the percentage of dead bee larvae in their least polluted sites was 10–30%, and, in contaminated sites, 50–60%. In pollen samples, we noted a higher Pb content, similar to Cd, and a much lower for Zn content than Moroń et al. [21]. The mortality of larvae in our study was similar to that found by Moroń et al. [21] on their least polluted sites.

Across all types of habitats, were found the same three species of parasites fly Cacoxenus indagator, the hymenopteran Monedontomerus obscurus and the mite Chaetodactylus osmiae. Of all the chambers built by O. bicornis females, parasites occupied 4.9% in berry plantations, 7.7% in urban habitat and 10.5% in orchards. We spotted no significant differences in the mean number of parasite chambers per nesting tube across the three habitat types (Table 1). Moroń et al. in 2014 [32] found losses caused by the same species of parasites at a level of approximately 1%. These are much lower values than those obtained in our study.

3.5. Implications of Using Osmia bicornis as Biomonitorsg

O. bicornis provides several reliable indicators—bee bodies, pollen, cocoons [66] and soil and leaves [35]. Moreover, the flight range of O. bicornis around the nest is approximately 600 m [36], and often even 100–200 m during the intensive flowering of plants. Each bee (female) is a forager, visiting flowers within a small area. All bees from one nest are exposed to similar pollutants. This makes O. bicornis a competitor to the honey bee. As reported by Bogdanov (2006) [67], the levels of metals in honey are lower than in the bees’ bodies, which means that bees can filter and purify the nectar to remove these agents. Borsuk et al., (2021) [64] confirmed that bees purify nectar by removing metals, as previously proposed by Bogdanov (2006) [68] and Dzhugan et al., (2018) [68]. Silici et al., (2016), also concluded that honey is not a good indicator of environmental pollution [59]. Conti and Botrè 2001 claimed that only pollen, propolis, wax, and the bees themselves are useful for assessing the content of heavy metals in the environment [56]. O. bicornis provides two of the mentioned indicators—bee bodies and pollen. Homogenized or bulk honey bee samples have been used for decades to monitor metal pollution in the environment. When using honeybees as bioindicators, it is advisable to sample more beehives at each location to ensure that there is an appropriate data set for a reliable interpretation of the results [48] however this increases the cost of monitoring. Some of the honey bees samples may be collectors, while others may be feeding bees, and each of these castes may have a different elemental load. “Colecting bees” that are from the same hive may forage in different places and have different exposures to pollutants. Foragers are the only type that come out the hive and accumulate pollutants [69]. De Vere et al., (2017) [70] and Dimou and Thrasyvoulou (2007) [71] showed that honeybees from one apiary collect pollen from different sources. Common honey bee foragers fly within a 1.5 km radius of the hive, visiting up to 10 million flowers per day [72]. These bees may be exposed to toxic substances to varying degrees and at different distances from the hive. Many of the above studies as well as our results indicate that O. bicornis can be used as a bioindicator.

4. Conclusions

Summarizing our results, we found no significant discrepancies in heavy-metal concentrations across the berry plantations, orchards or urban habitats. Habitat type had no effect on most bee reproductive parameters (percentage of occupied nest tubes, population growth). One can conclude, however, that considerably fewer bee larvae died in orchards than in berry plantations. Our study shows that bees that pollinate crops such as berries or those in orchards are exposed to heavy metals to a similar extent as in urban areas. Our research confirms that O. bicornis bees can be used as a bioindicator in various types of habitats.