DOI: 10.5061/dryad.v41ns1rxz
title: Data for: Termites have wider thermal limits to cope with environmental conditions in savannas

description: The most diverse and abundant family of termites, the Termitidae, evolved
 in African tropical forests. They have since colonised grassy biomes such
 as savannas. These open environments have more extreme conditions than
 tropical forests, notably wider extremes of temperature and lower
 precipitation levels and greater temporal fluctuations (both annual and
 diurnal variation). These conditions are challenging for soft-bodied
 ectotherms, such as termites, to survive in, let alone become as
 ecologically dominant as termites have. Here, we quantified termite
 thermal limits to test the hypothesis that these physiological limits have
 widened in savanna termite species to facilitate their existence in
 savanna environments. We sampled termites directly from mound structures,
 across an environmental gradient in Ghana, ranging from wet tropical
 forest through to savanna. At each location we quantified both Critical
 Thermal Maximum (CTmax) and Critical Thermal Minimum (CTmin) of all the
 most abundant mound-building Termitidae species in the study areas. We
 modelled the thermal limits in two separate mixed effects models against:
 canopy cover at the mound, temperature and rainfall, as fixed effects,
 with sampling location as a random intercept. For both CTmax and CTmin
 savanna species had significantly more extreme thermal limits than forest
 species. Between and within environments, areas with higher amounts of
 canopy cover were significantly associated with lower CTmax values of the
 termite colonies. CTmin was significantly positively correlated with
 rainfall. Temperature was retained in both models, however it did not have
 a significant relationship in either.  Sampling location
 explained a large proportion of the residual variation, suggesting there
 are other environmental factors that could influence termite thermal
 limits. Our results suggest there has been a widening of the thermal
 limits in termite savanna species. These physiological differences, in
 conjunction with other behavioural adaptations, are likely to have enabled
 termites to cope with the more extreme environmental conditions found in
 savanna environments and facilitated their expansion into open tropical
 environments.
description: Termites were sampled from
 four locations in Ghana across two different biomes: tropical rainforest
 and Guinea savanna. The three forest locations were: Bobiri Forest
 Reserve, a moist semi-deciduous forest in the Ashanti Region, a matrix of
 forest patches that have been, or currently are, logged (all colonies
 sampled from fragments that have not been logged for at least 20 years);
 the Forestry Research Institute of Ghana (FoRIG) Campus, a moist
 semi-deciduous forest patch located on the campus at Fumesua, near Kumasi,
 in the Ashanti Region; and the Assin-Attandanso Resource Reserve side of
 the Kakum Conservation Area, a moist evergreen tropical forest that has
 not been logged since 1990 (Wiafe, 2016), in the Central Region. Termites
 were sampled from a single savanna site, Mole National Park, a large
 unfenced protected area in the Savannah Region, between June and July
 2019. Sampling in all locations was conducted during the end of the dry
 season and the start of the wet season. Termites were sampled during the
 end of the dry season and start of the wet season at all sites: between
 January and March 2019 for the three forest sites, and June and July 2019
 for the savanna site. <strong>Termite and
 Environmental Sampling</strong> We sampled termites directly from
 termite mounds, identified to genus level (based on their diagnostic mound
 structures), which we located using directed searching, using the
 knowledge and experience of the field assistants. Two mounds were sampled
 per day, typically between 07h00 and 09h00. Target mounds were broken open
 and we collected termites, as well as some mound material, and placed them
 into a cool box to protect the termites from external conditions and
 reduce stress that could affect their thermal tolerances while being
 transported to the laboratory. The sample from each colony was placed in a
 separate container to prevent interaction between the two colonies. Where
 possible, both worker and soldier castes were sampled. The time between
 sampling and experimentation was reduced as much as possible (mean = 122.6
 mins, sd = ±81.4 min). The large standard deviation is attributed to
 having a single collection, but running two experiments, per day. The same
 colonies that were sampled were used in two experiments, one testing
 thermal maximum and one testing thermal minimum (the order of which was
 decided randomly); hence, the individuals used in the second experiment
 were kept inside the cool box (which was out of direct sunlight) while the
 first experiment took place. We photographed each mound to
 assist with species identification. Canopy cover above each sampled mound
 was quantified using a hemispherical fish-eye lens (Canon EOS 70D camera,
 Sigma 4.5mm f/1:2.8 Circular Fisheye Lens). For large mounds the image was
 taken directly above the apex of the mound, but with small mounds
 (&lt;75 cm in height) the image was taken above the mound at a height
 of 75 cm due to the height of the stabilising monopod. Due to variable
 light conditions when the images were taken, we did not standardise ISO,
 shutter-speed or aperture, although variation in these settings were
 reduced as much as possible. Variable light conditions and camera settings
 were corrected using Gap Light Analyzer software (GLA; Frazer et al.,
 1999). The approximate leaf-area index above each mound was then
 calculated as a measure of canopy cover. Leaf-area index is a measurement
 of the quantity of leaf area in a canopy per unit area of ground, and
 ranges from 0 (no leaf cover) to 6 (complete leaf cover). <strong>Thermal Tolerance
 Experiments</strong> We used similar methods to those
 outlined in Bishop et al. (2017) to estimate the thermal tolerance of the
 termites. Fourteen individuals from each of two colonies (28 in total)
 were tested per experimental run. Only individuals of the helper castes,
 workers and soldiers, were used in the experiments Each termite was placed
 into a separate microcentrifuge tube. The tubes were then placed into a
 dry heat bath (Tropicooler
 260014‐2, Boekel Scientific, Feasterville, PA, USA; temperature range
 -19°C to 69°C and an accuracy of ±1°C). The heat bath contained two
 aluminium inserts with each consisting of 14 wells, each of which held a
 single microcentrifuge tube. Individuals from two colonies and different
 castes were randomly allocated to different wells to remove potential
 equipment bias. The two colonies sampled each day were used
 in each experimental run (CT<sub>max</sub> and
 CT<sub>min</sub>) and each colony was tested for both
 CT<sub>max</sub> and CT<sub>min</sub>; however,
 each individual termite was only used in one experiment. The temperature within each
 microcentrifuge tube was acclimated for 15 minutes.
 CT<sub>max</sub> experiments had an acclimation temperature of
 30°C, and
 CT<sub>min</sub> had an acclimation temperature of 24°C. After
 the acclimation period, the temperature was changed by 1°C (raised for a
 CT<sub>max</sub> experiment, and lowered for
 CT<sub>min</sub>), and maintained at each new temperature for
 a further three minutes. Following this three-minute period, we checked
 each termite for the onset of a heat or chill coma. Heat coma onset was
 considered to have occurred when the individual lost muscle co-ordination;
 if this occurred it was recorded as the CT<sub>max</sub> of
 that termite. A chill coma was considered to have occurred when there was
 a complete lack of movement despite flicking the microcentrifuge tube; if
 this occurred it was recorded as the CT<sub>min</sub> of that
 termite. When we noticed the termite had lost muscle co-ordination
 (CT<sub>max</sub>) or had stopped moving
 (CT<sub>min</sub>) the temperature was recorded as the thermal
 limit of that individual termite. The experimental run was ended when the
 thermal limit of all 28 termites was recorded.
 <strong>Termite
 Identification</strong> Termite colonies were
 identified to genus level in the field based on their mound structure. All
 termites used in experimental runs were preserved in 70% ethanol and
 brought to the UK. We took an individual from each colony and DNA was
 extracted from each sample. The <em>COII</em> mitochondrial
 gene was amplified and sequenced using Sanger Sequencing at the Natural
 History Museum, London. Each sequence was matched with the most closely
 related sequence on NCBI using BLAST (Johnson et al., 2008). If the
 sequences had a unique high likelihood identity match (&gt;98%) with a
 single species described on the NCBI database, that sample, and therefore
 the colony it was sampled from, was considered identified as that species.
 If there was not a unique high likelihood identity match, the sequences
 (and therefore the colonies from which each sample was taken) were grouped
 and labelled as an unspecified species within a genus (for example,
 <em>Cubitermes sp. A</em><em>)</em>. After
 identification, species that did not have at least three replicates per
 sampling location were removed from the dataset (19 colonies removed,
 present in online dataset). Since the genetic identification was
 conducted, the genus <em>Cubitermes</em> was split into
 several genera (Hellemans et al., 2020). However, there were limited
 sequence data for the new classifications so we have treated all the new
 genera as <em>Cubitermes sensu lato</em>.
description: Metadata tabs added in the datasets provide information as to the
 columns, their data type, units required, data format and a general
 description, which should provide the users with the necessary information
 to use these data.