1. INTRODUCTION
Urbanization is growing rapidly worldwide leading to habitat loss and modification, with detrimental consequences for biodiversity and ecological functioning (Ayers & Rehan, 2021; Wenzel et al. 2020; Fisogni et al., 2020). Wild pollinator insect communities are deeply affected by this phenomenon, especially in the case of bees (Hymenoptera: Anthophila). While some species are able to successfully exploit these conditions and thrive in urban areas (Fitch et al., 2019), others are facing challenges due to the lack of nesting and foraging habitat (Cane et al., 2005), distance between green areas and hostile climatic conditions (e.g. urban heat islands) (Biella et al., 2022), pollution (Tommasi et al., 2022a) or parasite infections (Youngsteadt et al., 2015). From a landscape perspective, urbanization significantly shapes land-use features such as composition (the proportion of different land use categories) and configuration (the spatial arrangement of patches of land-use categories) (Fu & Weng, 2016), with direct consequences for bees communities. In this context, green habitats act as islands which are exploited by wild bees, and whose loss has been associated to negative effects on bee species richness (Winfree et al., 2009) and their foraging habits (Tommasi et al., 2022b, Andrieu et al., 2009). Similarly, green habitat fragmentation may affect pollinator species foraging behavior (Andrieu et al., 2009) and richness with stronger consequences for the smaller and less mobile species (Ayers & Rehan, 2021).
Urbanization may also have cascading effects on the health and fitness of wild bees, for example by shaping the dynamics of their parasite infections (Figueroa et al., 2020) contributing to population decline of both managed and wild bee populations (Ivers et al., 2022). For instance, urban green habitat availability and fragmentation may indirectly affect parasite spread by shaping features of the host community such as species richness and abundance (Tommasi et al., 2022). Indeed, previous research focused on bumblebees suggested that urbanization may promote the diffusion of these parasites (e.g.,Crithidia spp. and Nosema spp. ) in wild bees (Theodorou et al., 2016) also due to changes in bumblebee community features (Ivers et al., 2022), increasing the pressure experienced by these organisms in urban habitat (but see Samuelson et al., 2020). However, despite the increasingly higher research effort towards parasite spillover among bees, the role of landscape and local features of green habitat in shaping parasite dynamics is a topic largely neglected in literature.
Inter and intraspecific transmission of parasites usually occur through direct contact between hosts or via contamination of flowers visited by infected individuals (Singh et al., 2010). In this context, a major role of beekeeping in contributing to the spread of parasites potentially infecting wild species has also been reported. (Dolezal et al., 2016 Meeus et al., 2011; Cilia et al., 2023). Since apiculture exceptionally increases the number of potential hosts in the area surrounding honeybee hives, this anthropic activity could facilitate both the direct contact between honeybees and wild species and the contamination of floral resources. Beekeeping has gained growing importance worldwide, and especially in urban areas mainly (Matsuzawa & Kohsaka, 2021), due to the positive impact on community building and environmental education (Skelton, 2006). Although several studies showed important effects of apiculture on parasite distribution in the wild pollinators community (Graystock et al., 2016a), nowadays dynamic and direct causes of this impact are still unclear (Cilia et al., 2023).
To investigate the role of environmental features on parasite occurrence different methodological approaches can be employed. For example histopathological evaluation as well as species-specific fluorescence in situ hybridization (FISH) can be employed for this purpose and have the advantages of being able to clearly localize the infections and also highlight the damages induced on tissues by parasites (Panek et al., 2018). Other methods such as those based on PCR can be efficient alternatives to more traditional approaches. Indeed, PCR-based methodologies are rapid and can be widely performed. Even if PCR detection does not allow discrimination between infection and contamination by non germinated spores present in the digestive tract (Gisder et al., 2020), it can reliably detect even low intensity or latent infections (Graystock et al., 2015). Hence, PCR methods are suitable for screening and could provide detailed insights on the role of urbanization in pollinator epidemiology and on pollinator-parasite interactions (Cohen el al., 2022.).
In this study we used a molecular approach to investigate the impact of urban green areas in shaping the occurrence of parasites in two bumblebee species, namely Bombus terrestris (Linnaeus, 1758) andB. pascuorum (Scopoli, 1763) both largely abundant also in cities (Tommasi et al., 2022). We focused on the parasite richness, indicating the number of different parasite taxa detected in each sample, and on the probability of infection thus of detecting at least a single parasite in a sample. Even if related to each other, these two variables indicate different aspects: while one indicates the possibility of co-infection and the parasite load, the other describes how probable it is to be infected by any of the studied parasite species. We considered several urban scales by focusing on the landscape structure, local features and honeybee presence as potential intermediate vectors. Specifically, we evaluated the impact of these features in shaping the occurrence of the commonest bumblebees parasites, trypanosomatids (Crithidia spp.), microsporidians (Nosema spp.) and neogregarines (Apicystis spp.) (Ivers et al., 2022; Theodorou et al., 2016; Youngstead et al., 2015). Since the reduced and fragmented green habitats of the more urbanized landscape are expected to concentrate bees to the remnant patches available for foraging (Dylewski et al., 2019; Quistberg et al., 2016), we hypothesized to observe an increase in the parasite richness and probability of infection in the smaller and more fragmented green habitats of the more urbanized areas. At the local scale, high availability and diversity of floral resources are expected to increase pollinator community richness and abundance (Tommasi et al., 2021, Hulsmann et al., 2015) thus following disease ecology theory (Becker et al.,, 2015) we hypothesized to observe also a positive correlation among flower abundance and parasite richness and probability of infection. Furthermore, based on previous research showing a positive correlation between beekeeping and the spread of parasites to wild species (Graystock et al., 2016a), we expected a higher parasite richness and probability of infection in bumblebees collected from study sites with a higher abundance of honeybee hives in the surrounding and/or located closer to the apiary.