Introduction
The concept of extended phenotype lies in the idea that a gene exerts
effects beyond the organism itself, by creating physical structures,
modifying the environment or manipulating other organisms to increase
its reproductive success. Ever since Dawkins introduced the theoretical
framework of “extended phenotype” (Dawkins, 1982), contemporary
evolutionary biology has demonstrated its influence by providing
numerous examples of complex relationships and adaptations in all
taxonomic groups (Camargo & Pedro, 2003; Whitham et al., 2003; Cui et
al., 2020; Fernandez et al., 2022; DeLong et al., 2022). As the
introduction of this conceptual framework stimulated both heated debates
and a great deal of research activity (Dawkins, 2004; Hunter, 2018), its
incorporation into quantitative and empirical approaches in an attempt
to bridge the gap between genotypes and environments towards an
integrated theory of evolution remains a challenge. The environment,
usually seen as an independent component of phenotypic variation in
quantitative genetics, is instead viewed as a variable affected by the
genotype (Bailey, 2012; Edelaar et al., 2023). The investigation into
extended phenotypes, including behavioral manipulation or housing
structure and associated microorganisms, has many potential applications
in the fields of ecology and agriculture (Woods et al., 2021; Favela et
al., 2023). This is particularly relevant for keystones species (“whose
impact on its community or ecosystem is large, and disproportionately
large relative to its abundance” (Power et al., 1996)) such as social
insects (Elizalde et al., 2020). In the case of social bees, their
ecological and economical importance triggered research on their related
extended phenotype, mainly focusing on nesting (Smith et al., 2015),
foraging and communication (Hughes, 2008; Schaedelin & Taborsky, 2009)
yet overlooking their most renowned product, honey. Honey production
combines the features of the extended phenotype with a well-defined
genetic basis, from foraging (Page et al., 2000) to nectar collection
and processing by bee enzymes (Kunieda et al., 2006; Vannette et al.,
2015) and symbiotic microorganisms (de Paula et al., 2021). Further,
this contributes to the colony fitness not only as stored source of
carbohydrates, but also for improving immunity and regulating
temperature, development and adult longevity (Berenbaum & Calla, 2021).
Although the production of honey has been the hallmark of Apis
mellifera in Europe, the well-known Western honey bee spread
across the entire world, another tribe of social bees called stingless
bees (Apidae, Subfamily Meliponinae) are found exclusively in the
(sub-)tropical regions and produce honey used for its nutritional and
medicinal properties (Chuttong et al., 2018; Pimentel et al., 2021;
Noiset et al., 2022; Héger et al., 2023; Vit et al., 2024).
Approximately 600 species of stingless bees belonging to 45 genera have
been described to this date (Engel et al., 2023), making Meliponinae the
largest group of corbiculate bees. Despite this great diversity,
stingless bees and their products have been relatively less studied than
honey bees and have mainly been investigated in a few Neotropical
countries (Brazil & Mexico in the lead), neglecting other Neotropical
countries, the Afrotropics and the Indomalayan, Papuasian, and
Australian (IPA) region (Nordin et al., 2018). Stingless bee honey (SBH)
has higher water content than honey from A. mellifera , which
allows the persistence of hydrophilous symbiotic microorganisms involved
in sugar fermentation (Menezes et al., 2013). This is a key process for
the preservation of honey that results in a less sweet and more acidic
product (Biluca et al., 2016) but also in the production of a range of
unique by-products associated with health benefits (Fletcher et al.,
2020). Additionally, honey microbiota and phytochemicals transferred
from honey pots made of cerumen (a mix of wax and propolis) contributes
to its medicinal properties by introducing antimicrobial compounds to
reduce the proliferation of competing microorganism (Ngalimat et al.,
2020; Chuttong et al., 2023). Therefore, SBH may be the extended
phenotype of the hive but also the one of the microorganisms hosted by
honey and bees. These microorganisms benefits as well from this aqueous
solutions that has been proposed to be the natural habitat of some
osmophilic yeasts (Matos et al., 2020).
The understanding of the compositional variability of SBH is still
limited due to the numerous sources of variation interacting with each
other. Biochemical reactions in stingless bee honey vary according to
the bee species producing enzymes, processing dynamics and the
microbiota (Souza et al., 2021). Honey microbiota varies according to
the foraging patterns, nectar microbiota, environment, hive and the bee
species trough vertical and horizontal transmission, the relative
importance of each has yet to be defined (Cerqueira et al., 2024).
Stingless bee also visits a wide variety of (sub-)tropical plants (Bueno
et al., 2023) whose nectar and microbiota composition vary, adding
significant levels of phytochemicals and amino acids to honey (Shamsudin
et al., 2019; Biluca et al., 2019). Nectar composition itself is
influenced by ecological (water availability, temperature, soil factors,
interactions with herbivores and nectar robbers) and evolutionary
drivers such as plant phylogenetic constraints and pollination syndrome
(Nicolson & Thornburg, 2007; Parachnowitsch et al., 2019). Foraging
activity of stingless bees also include resin collection to build nests
and honey pots which also influence honey composition and properties
(Kegode et al., 2023; Villagomez et al., 2024; Nakamura et al., 2024).
Beyond these complex factors, the lack of scientific research on the
physico-chemical characteristics and bio-functional properties of SBH
from the Afrotropics and the Indomalayan, Papuasian, and Australian
regions hinder the exploration of the evolutionary ecology hypothesis
regarding SBH.
To fill these gaps, our study aimed to work at the global level to i)
characterize the differences between the composition of honeys from
stingless bees and honey bees using advanced chemometric methods, ii)
investigate patterns of compositional variation among tropical regions
and finally iii) disentangle the roles of evolutionary history and
environmental conditions on honey properties. Ultimately, we aim to
provide the basis of a theoretical framework to understand the drivers
of compositional variability of SBH.