Genomic evidence for three distinct species in the Erebia manto complex in Central Europe (Lepidoptera, Nymphalidae).

A problem to implement conservation strategies is that in many cases recognized taxa are in fact complexes of several cryptic species. Failure to properly delineate species may lead to misplaced priorities or to inadequate conservation measures. One such species complex is the yellow-spotted ringlet Erebia manto, which comprises several phenotypically distinct lineages, whose degree of genomic isolation has so far not been assessed. Some of these lineages are geographically restricted and thus possibly represent distinct units with conservation priorities. Using several thousand nuclear genomic markers, we evaluated to which degree the bubastis lineage from the Alps and the vogesiaca lineage from the Vosges, are genetically isolated from the widespread manto lineage. Our results suggest that both lineages are genetically as strongly differentiated from manto as other taxonomically well separated sibling species in this genus from each other, supporting a delineation of bubastis and vogesiaca as independent species. Given the restricted and isolated range of vogesiaca as well as the disjunct distribution of bubastis, our findings have significant implication for future conservation efforts on these formerly cryptic species and highlight the need to investigate the genomic identity within species complexes. Supplementary Information: The online version contains supplementary material available at 10.1007/s10592-023-01501-w.

The new annotated checklist of the wild bees of Europe (Hymenoptera: Anthophila).

At a time when nature conservation has become essential to ensure the long-term sustainability of our environment, it is widely acknowledged that conservation actions must be implemented within a solid taxonomic framework. In preparation for the upcoming update of the IUCN Red List, we here update the European checklist of the wild bees (sensu the IUCN geographical framework). The original checklist, published in 2014, was revised for the first time in 2017. In the present revision, we add one genus, four subgenera and 67 species recently described, 40 species newly recorded since the latest revision (including two species that are not native to Europe), 26 species overlooked in the previous European checklists and 63 published synonymies. We provide original records for eight species previously unknown to the continent and, as original taxonomic acts, we provide three new synonyms, we consider two names as nomina nuda, ten names as nomina dubia, three as species inquirenda, synonymize three species and exclude 40 species from the previous checklist. Around a hundred other taxonomic changes and clarifications are also included and discussed. The present work revises the total number of genera for IUCN Europe to 77 and the total number of species to 2,138. In addition to specifying the taxonomic changes necessary to update the forthcoming Red List of European bees, we discuss the sampling and taxonomic biases that characterise research on the European bee fauna and highlight the growing importance of range expansions and species invasions.

Taxonomic revision of the subgenus Eutricharaea Thomson in Egypt, with a key to the species and the description of two new species (Hymenoptera, Anthophila, Megachilidae, genus Megachile Latreille).

The subgenus Eutricharaea Thomson of the genus Megachile Latreille (Hymenoptera: Megachilidae) in Egypt is revised. Fifteen species are recognized, of which two are new: Megachile laniventris Praz sp. nov. and M. rufomandibularis Praz sp. nov. In addition, three unclear taxa known from few specimens in one sex are documented to facilitate future work. The following new synonymies are proposed: M. microxantha Cockerell 1937, M. privigna Rebmann 1968, and M. submucida Alfken 1926 are synonymized with M. leucostoma Prez 1907 (syn. nov.); M. blanda Rebmann 1968 with M. walkeri Dalla Torre 1896 (syn. nov.); M. uniformis Alfken 1934 (not M. uniformis Mitchell 1929) and the replacement name M. minutuloides Alfken 1936 with M. minutissima Radoszkowski 1876 (syn. nov.); and M. tkalcui Zanden 1996 with M. rugipuncta Alfken 1934 (syn. nov.). Lectotypes are designated for M. soikai Benoist 1961, M. rugipuncta, M. tenuistriga Alfken 1938, M. niveascopa Ferton 1908, and M. naevia Kohl 1906. The previously unknown males of M. impressipuncta Alfken 1934 and M. rugipuncta, as well as the undescribed female of M. soikai are described. Illustrated keys for both males and females are provided and DNA barcodes are published for most species.

Global patterns in bumble bee pollen collection show phylogenetic conservation of diet.

Bumble bees (Bombus) are a group of eusocial bees with a strongly generalised feeding pattern, collecting pollen from many different botanical families. Though predominantly generalists, some bumble bee species seem to have restricted dietary choices. It is unclear whether restricted diets in bumble bees are inherent or a function of local conditions due to a lack of data for many species across different regions. The objective of this study was to determine whether bumble bee species displayed specific patterns of pollen collection, and whether patterns were influenced by phylogenetic relatedness or tongue length, a trait known to be associated with structuring floral visitation. Bumble bee pollen collection patterns were quantified from 4,132 pollen loads taken from 58 bumble bee species, representing 24% of the pollen-collecting diversity of this genus. Phylogenetic trait mapping showed a conserved pattern of dietary dissimilarity across species, but not for dietary breadth. Dietary dissimilarity was driven by collection of Fabaceae, with the most similar species collecting around 50%-60% of their diet from this botanical family. The proportion of the diet collected from Fabaceae also showed a conserved phylogenetic signal. Greater collection of Fabaceae was associated with longer tongue lengths, with shorter tongued species focusing on alternative botanical families. However, this result was largely driven by phylogenetic relatedness, not tongue length per se. These results demonstrate that, though generalists, bumble bees are still subject to dietary restrictions that constrain their foraging choices. These dietary constraints have implications for their persistence should their core resources decline in abundance.

The wild bees (Hymenoptera: Apoidea) of Morocco.

Morocco is a well known hot-spot of biodiversity in the Mediterranean basin. While some taxa like vascular plants are relatively well recorded, important groups of pollinators like bees are still understudied. This article presents an updated checklist of the bee species of Morocco and includes a summary of global and regional distribution of each species. A total of 961 species belonging to six bee families and 68 genera are recorded: Andrenidae (8 genera, 217 species); Apidae (15 genera, 241 species); Colletidae (2 genera, 74 species), Halictidae (12 genera, 144 species), Megachilidae (28 genera, 271 species) and Melittidae (3 genera, 14 species). Among them, 67 species are recorded for the first time in Morocco. Around 70% of the bee fauna of Morocco consists of widespread Palaearctic species. Only 18% of Moroccan species recorded are restricted to North Africa and 8% are Moroccan single-country endemics (81 species). Afrotropical elements in the Moroccan fauna are few, with only 3% of Morocco species co-occuring in that region. This checklist is intended to stimulate new regional research on bees including their taxonomy and biogeography. As many groups of bees have been understudied, discovery of new species for science and new records for the country can be expected. Additional research including inventorying, monitoring, and integrative taxonomic studies are needed to develop a comprehensive strategy for bee conservation in Morocco.

Ultraconserved yet informative for species delimitation: Ultraconserved elements resolve long-standing systematic enigma in Central European bees.

Accurate and testable species hypotheses are essential for measuring, surveying and managing biodiversity. Taxonomists often rely on mitochondrial DNA barcoding to complement morphological species delimitations. Although COI-barcoding has largely proven successful in assisting identifications for most animal taxa, there are nevertheless numerous cases where mitochondrial barcodes do not reflect species hypotheses. For instance, what is regarded as a single species can be associated with two distinct DNA barcodes, which can point either to cryptic diversity or to within-species mitochondrial divergences without reproductive isolation. In contrast, two or more species can share barcodes, for instance due to mitochondrial introgression. These intrinsic limitations of DNA barcoding are commonly addressed with nuclear genomic markers, which are expensive, may have low repeatability and often require high-quality DNA. To overcome these limitations, we examined the use of ultraconserved elements (UCEs) as a quick and robust genomic approach to address such problematic cases of species delimitation in bees. This genomic method was assessed using six different species complexes suspected to harbour cryptic diversity, mitochondrial introgression or mitochondrial paraphyly. The sequencing of UCEs recovered between 686 and 1,860 homologous nuclear loci and provided explicit species delimitation in all investigated species complexes. These results provide strong evidence for the suitability of UCEs as a fast method for species delimitation even in recently diverged lineages. Furthermore, we provide the first evidence for both mitochondrial introgression among distinct bee species, and mitochondrial paraphyly within a single bee species.

Evaluating next-generation sequencing (NGS) methods for routine monitoring of wild bees: Metabarcoding, mitogenomics or NGS barcoding.

Implementing cost-effective monitoring programs for wild bees remains challenging due to the high costs of sampling and specimen identification. To reduce costs, next-generation sequencing (NGS)-based methods have lately been suggested as alternatives to morphology-based identifications. To provide a comprehensive presentation of the advantages and weaknesses of different NGS-based identification methods, we assessed three of the most promising ones, namely metabarcoding, mitogenomics and NGS barcoding. Using a regular monitoring data set (723 specimens identified using morphology), we found that NGS barcoding performed best for both species presence/absence and abundance data, producing only few false positives (3.4%) and no false negatives. In contrast, the proportion of false positives and false negatives was higher using metabarcoding and mitogenomics. Although strong correlations were found between biomass and read numbers, abundance estimates significantly skewed the communities' composition in these two techniques. NGS barcoding recovered the same ecological patterns as morphology. Ecological conclusions based on metabarcoding and mitogenomics were similar to those based on morphology when using presence/absence data, but different when using abundance data. In terms of workload and cost, we show that metabarcoding and NGS barcoding can compete with morphology, but not mitogenomics which was consistently more expensive. Based on these results, we advocate that NGS barcoding is currently the seemliest NGS method for monitoring of wild bees. Furthermore, this method has the advantage of potentially linking DNA sequences with preserved voucher specimens, which enable morphological re-examination and will thus produce verifiable records which can be fed into faunistic databases.

A DNA barcode reference library for Swiss butterflies and forester moths as a tool for species identification, systematics and conservation.

Butterfly monitoring and Red List programs in Switzerland rely on a combination of observations and collection records to document changes in species distributions through time. While most butterflies can be identified using morphology, some taxa remain challenging, making it difficult to accurately map their distributions and develop appropriate conservation measures. In this paper, we explore the use of the DNA barcode (a fragment of the mitochondrial gene COI) as a tool for the identification of Swiss butterflies and forester moths (Rhopalocera and Zygaenidae). We present a national DNA barcode reference library including 868 sequences representing 217 out of 224 resident species, or 96.9% of Swiss fauna. DNA barcodes were diagnostic for nearly 90% of Swiss species. The remaining 10% represent cases of para- and polyphyly likely involving introgression or incomplete lineage sorting among closely related taxa. We demonstrate that integrative taxonomic methods incorporating a combination of morphological and genetic techniques result in a rate of species identification of over 96% in females and over 98% in males, higher than either morphology or DNA barcodes alone. We explore the use of the DNA barcode for exploring boundaries among taxa, understanding the geographical distribution of cryptic diversity and evaluating the status of purportedly endemic taxa. Finally, we discuss how DNA barcodes may be used to improve field practices and ultimately enhance conservation strategies.

Taxonomic revision of the Western Palaearctic bees of the subgenus Pseudomegachile (Hymenoptera, Apiformes, Megachilidae, Megachile).

The subgenus Pseudomegachile Friese of the large genus Megachile Latreille is revised for the Western Palaearctic region, Iran, and the Arabian Peninsula. Twenty species are recognized, of which five are new: Megachile blepharis Dorchin Praz from Israel, M. plumigera Dorchin Praz from Oman and the United Arab Emirates, M. syriaca Dorchin Praz from Syria and Turkey, M. yezidica Dorchin Praz from Turkey and Iran, and M. maxschwarzi Dorchin Praz from Iran and Central Asia. The following new synonymies are proposed: M. inermis Radoszkowski 1893 (as well as the replacement name M. mitis Cockerell 1899), M. albifasciata Rebmann 1970 and M. transgrediens Rebmann 1970 are placed in synonymy with M. saussurei Radoszkowski 1874; M. tuberculata Radoszkowski 1893 (as well as the replacement name M. tuberculosa Dalla Torre 1896) is placed in synonymy with M. seraxensis Radoszkowski 1893; M. rubripes Morawitz 1875 and M. persica Rebmann 1972 are placed in synonymy with M. flavipes Spinola 1838; M. stolzmanni Radoszkowski 1893 and M. flavidula Rebmann 1970 are placed in synonymy with M. tecta Radoszkowski 1888. Lectotypes are designated for M. cinnamomea Alfken 1926, M. nilotica Pérez 1897, M. inermis, M. seraxensis, M. tuberculata, M. farinosa Smith 1853, M. derasa Gerstäcker 1869, M. erythrocnemis Alfken 1930 and M. xanthocnemis Alfken 1938. An identification key is provided, a phylogenetic hypothesis including most species in the subgenus is presented, and the biology of the species is briefly summarized.

Nursing protects honeybee larvae from secondary metabolites of pollen.

The pollen of many plants contains toxic secondary compounds, sometimes in concentrations higher than those found in the flowers or leaves. The ecological significance of these compounds remains unclear, and their impact on bees is largely unexplored. Here, we studied the impact of pyrrolizidine alkaloids (PAs) found in the pollen of Echium vulgare on honeybee adults and larvae. Echimidine, a PA present in E. vulgare pollen, was isolated and added to the honeybee diets in order to perform toxicity bioassays. While adult bees showed relatively high tolerance to PAs, larvae were much more sensitive. In contrast to other bees, the honeybee larval diet typically contains only traces of pollen and consists predominantly of hypopharyngeal and mandibular secretions produced by nurse bees, which feed on large quantities of pollen-containing bee bread. We quantified the transfer of PAs to nursing secretions produced by bees that had previously consumed bee bread supplemented with PAs. The PA concentration in these secretions was reduced by three orders of magnitude as compared to the PA content in the nurse diet and was well below the toxicity threshold for larvae. Our results suggest that larval nursing protects honeybee larvae from the toxic effect of secondary metabolites of pollen.

Pyrrolizidine Alkaloids from Echium vulgare in Honey Originate Primarily from Floral Nectar.

Pyrrolizidine alkaloids (PAs) in honey can be a potential human health risk. So far, it has remained unclear whether PAs in honey originate from pollen or floral nectar. We obtained honey, nectar, and plant pollen from two observation sites where Echium vulgare L. was naturally abundant. The PA concentration of honey was determined by targeted analysis using a high pressure liquid chromatography-mass spectrometry system (HPLC-MS/MS), allowing the quantification of six different PAs and PA-N-oxides present in E. vulgare. Echium-type PAs were detected up to 0.153 µg/g in honey. Nectar and plant pollen were analyzed by nontargeted analysis using ultrahigh pressure liquid chromatography-high resolution-mass spectrometry (UHPLC-HR-MS), allowing the detection of 10 alkaloids in small size samples. Echium-type PAs were detected between 0.3-95.1 µg/g in nectar and 500-35000 µg/g in plant pollen. The PA composition in nectar and plant pollen was compared to the composition in honey. Echimidine (+N-oxide) was the main alkaloid detected in honey and nectar samples, while echivulgarine (+N-oxide) was the main PA found in plant pollen. These results suggest that nectar contributes more significantly to PA contamination in honey than plant pollen.

Phylogenomics Controlling for Base Compositional Bias Reveals a Single Origin of Eusociality in Corbiculate Bees.

As increasingly large molecular data sets are collected for phylogenomics, the conflicting phylogenetic signal among gene trees poses challenges to resolve some difficult nodes of the Tree of Life. Among these nodes, the phylogenetic position of the honey bees (Apini) within the corbiculate bee group remains controversial, despite its considerable importance for understanding the emergence and maintenance of eusociality. Here, we show that this controversy stems in part from pervasive phylogenetic conflicts among GC-rich gene trees. GC-rich genes typically have a high nucleotidic heterogeneity among species, which can induce topological conflicts among gene trees. When retaining only the most GC-homogeneous genes or using a nonhomogeneous model of sequence evolution, our analyses reveal a monophyletic group of the three lineages with a eusocial lifestyle (honey bees, bumble bees, and stingless bees). These phylogenetic relationships strongly suggest a single origin of eusociality in the corbiculate bees, with no reversal to solitary living in this group. To accurately reconstruct other important evolutionary steps across the Tree of Life, we suggest removing GC-rich and GC-heterogeneous genes from large phylogenomic data sets. Interpreted as a consequence of genome-wide variations in recombination rates, this GC effect can affect all taxa featuring GC-biased gene conversion, which is common in eukaryotes.

Phylogenetic position of the bee genera Ancyla and Tarsalia (Hymenoptera: Apidae): a remarkable base compositional bias and an early Paleogene geodispersal from North America to the Old World.

We address the phylogenetic position of the bee genera Tarsalia and Ancyla (currently forming the tribe Ancylaini) on the basis of morphological, molecular and combined data. We assembled a matrix of 309 morphological characters and 5246 aligned nucleotide positions from six nuclear genes (28S, EF-1a, wingless, POL2, LW-Rhodopsin, NAK). In addition to both constituent genera of Ancylaini, we include all three subtribes of the Eucerini as well as a large number of other tribes from the "eucerine line". The morphological data suggest Ancyla to be sister to Tarsalia+Eucerini and analyses of the entire molecular dataset suggest Tarsalia to be sister to Ancyla+Eucerini. However, analyses of the combined dataset suggests the Ancylaini to be monophyletic. We address possible bias within the molecular data and show that the base composition of two markers (EF-1a and NAK) is significantly heterogeneous among taxa and that this heterogeneity is strong enough to overcome the phylogenetic signal from the other markers. Analyses of a molecular matrix where the heterogeneous partitions have been RY-recoded yield trees that are better resolved and have higher nodal support values than those recovered in analyses of the non-recoded matrix, and strongly suggest the Ancylaini to be a monophyletic sister group to the Eucerini. A dated phylogeny and ancestral range reconstructions suggest that the common ancestor of the Ancylaini reached the Old World from the New World most probably via the Thulean Land Bridge in a time window between 69 and 47 mya, a period that includes the Early Eocene Climatic Optimum. No further exchanges between the New World and the Old World are implied by our data until the period between 22 mya and 13.9 mya. These more recent faunal exchanges probably involved geodispersal over the Bering Land Bridge by less thermophilic lineages.

A new species of the paper wasp genus Polistes (Hymenoptera, Vespidae, Polistinae) in Europe revealed by morphometrics and molecular analyses.

We combine multivariate ratio analysis (MRA) of body measurements and analyses of mitochondrial and nuclear data to examine the status of several species of European paper wasps (Polistes Latreille, 1802) closely related to P. gallicus. Our analyses unambiguously reveal the presence of a cryptic species in Europe, as two distinct species can be recognized in what has hitherto been considered Polistes bischoffi Weyrauch, 1937. One species is almost as light coloured as P. gallicus, and is mainly recorded from Southern Europe and Western Asia. The other species is darker and has a more northern distribution in Central Europe. Both species occur syntopically in Switzerland. Given that the lost lectotype of P. bischoffi originated from Sardinia, we selected a female of the southern species as a neotype. The northern species is described as P. helveticus sp. n. here. We also provide a redescription of P. bischoffi rev. stat. and an identification key including three more closely related species, P. biglumis, P. gallicus and P. hellenicus.

Origins, evolution, and diversification of cleptoparasitic lineages in long-tongued bees.

The evolution of parasitic behavior may catalyze the exploitation of new ecological niches yet also binds the fate of a parasite to that of its host. It is thus not clear whether evolutionary transitions from free-living organism to parasite lead to increased or decreased rates of diversification. We explore the evolution of brood parasitism in long-tongued bees and find decreased rates of diversification in eight of 10 brood parasitic clades. We propose a pathway for the evolution of brood parasitic strategy and find that a strategy in which a closed host nest cell is parasitized and the host offspring is killed by the adult parasite represents an obligate first step in the appearance of a brood parasitic lineage; this ultimately evolves into a strategy in which an open host cell is parasitized and the host offspring is killed by a specialized larval instar. The transition to parasitizing open nest cells expanded the range of potential hosts for brood parasitic bees and played a fundamental role in the patterns of diversification seen in brood parasitic clades. We address the prevalence of brood parasitic lineages in certain families of bees and examine the evolution of brood parasitism in other groups of organisms.

The impact of molecular data on our understanding of bee phylogeny and evolution.

Our understanding of bee phylogeny has improved over the past fifteen years as a result of new data, primarily nucleotide sequence data, and new methods, primarily model-based methods of phylogeny reconstruction. Phylogenetic studies based on single or, more commonly, multilocus data sets have helped resolve the placement of bees within the superfamily Apoidea; the relationships among the seven families of bees; and the relationships among bee subfamilies, tribes, genera, and species. In addition, molecular phylogenies have played an important role in inferring evolutionary patterns and processes in bees. Phylogenies have provided the comparative framework for understanding the evolution of host-plant associations and pollen specialization, the evolution of social behavior, and the evolution of parasitism. In this paper, we present an overview of significant discoveries in bee phylogeny based primarily on the application of molecular data. We review the phylogenetic hypotheses family-by-family and then describe how the new phylogenetic insights have altered our understanding of bee biology.

Why do leafcutter bees cut leaves? New insights into the early evolution of bees.

Stark contrasts in clade species diversity are reported across the tree of life and are especially conspicuous when observed in closely related lineages. The explanation for such disparity has often been attributed to the evolution of key innovations that facilitate colonization of new ecological niches. The factors underlying diversification in bees remain poorly explored. Bees are thought to have originated from apoid wasps during the Mid-Cretaceous, a period that coincides with the appearance of angiosperm eudicot pollen grains in the fossil record. The reliance of bees on angiosperm pollen and their fundamental role as angiosperm pollinators have contributed to the idea that both groups may have undergone simultaneous radiations. We demonstrate that one key innovation--the inclusion of foreign material in nest construction--underlies both a massive range expansion and a significant increase in the rate of diversification within the second largest bee family, Megachilidae. Basal clades within the family are restricted to deserts and exhibit plesiomorphic features rarely observed among modern bees, but prevalent among apoid wasps. Our results suggest that early bees inherited a suite of behavioural traits that acted as powerful evolutionary constraints. While the transition to pollen as a larval food source opened an enormous ecological niche for the early bees, the exploitation of this niche and the subsequent diversification of bees only became possible after bees had evolved adaptations to overcome these constraints.

Phylogeny and biogeography of bees of the tribe Osmiini (Hymenoptera: Megachilidae).

The Osmiini (Megachilidae) constitute a taxonomically and biologically diverse tribe of bees. To resolve their generic and suprageneric relationships, we inferred a phylogeny based on three nuclear genes (Elongation factor 1-alpha, LW-rhodopsin and CAD) applying both parsimony and Bayesian methods. Our phylogeny, which includes 95 osmiine species representing 18 of the 19 currently recognized genera, is well resolved with high support for most basal nodes. The core osmiine genera were found to form a well-supported monophyletic group, but four small genera, Noteriades, Afroheriades,Pseudoheriades and possibly Ochreriades, formerly included in the Osmiini, do not appear to belong within this tribe. Our phylogeny results in the following taxonomic changes: Stenosmia and Hoplosmia are reduced to subgeneric rank in Hoplitis and Osmia, respectively, Micreriades is recognized as a subgenus in Hoplitis and the subgenus Nasutosmia is transferred from Hoplitis to Osmia. We inferred a biogeographic scenario for the Osmiini applying maximum likelihood inference and models of character evolution. We provide evidence that the Osmiini originated in the Palearctic, and that extensive exchanges occurred between the Palearctic and the Nearctic. The latter finding may relate to the fact that many osmiine species nest in wood or in stems, facilitating dispersal by overseas transport of the nests.

Patterns of host-plant choice in bees of the genus Chelostoma: the constraint hypothesis of host-range evolution in bees.

To trace the evolution of host-plant choice in bees of the genus Chelostoma (Megachilidae), we assessed the host plants of 35 Palearctic, North American and Indomalayan species by microscopically analyzing the pollen loads of 634 females and reconstructed their phylogenetic history based on four genes and a morphological dataset, applying both parsimony and Bayesian methods. All species except two were found to be strict pollen specialists at the level of plant family or genus. These oligolectic species together exploit the flowers of eight different plant orders that are distributed among all major angiosperm lineages. Based on ancestral state reconstruction, we found that oligolecty is the ancestral state in Chelostoma and that the two pollen generalists evolved from oligolectic ancestors. The distinct pattern of host broadening in these two polylectic species, the highly conserved floral specializations within the different clades, the exploitation of unrelated hosts with a striking floral similarity as well as a recent report on larval performance on nonhost pollen in two Chelostoma species clearly suggest that floral host choice is physiologically or neurologically constrained in bees of the genus Chelostoma. Based on this finding, we propose a new hypothesis on the evolution of host range in bees.

Specialized bees fail to develop on non-host pollen: do plants chemically protect their pollen?

Bees require large amounts of pollen for their own reproduction. While several morphological flower traits are known to have evolved to protect plants against excessive pollen harvesting by bees, little is known on how selection to minimize pollen loss acts on the chemical composition of pollen. In this study, we traced the larval development of four solitary bee species, each specialized on a different pollen source, when reared on non-host pollen by transferring unhatched eggs of one species onto the pollen provisions of another species. Pollen diets of Asteraceae and Ranunculus (Ranunculaceae) proved to be inadequate for all bee species tested except those specialized on these plants. Further, pollen of Sinapis (Brassicaceae) and Echium (Boraginaceae) failed to support larval development in one bee species specialized on Campanula (Campanulaceae). Our results strongly suggest that pollen of these four taxonomic groups possess protective properties that hamper digestion and thus challenge the general view of pollen as an easy-to-use protein source for flower visitors.