Short-term survival of incipient colonies
Fourteen days after pairing, only 101 incipient colonies (202 alates) of the 831 established pairings survived; 35 out of the 231 inbred pairings (15.15%) and 66 out of the 600 outbred pairings (11.00%). No significant difference was observed between the survival of inbred and outbred pairings (P = 0.212; Figure 1a). However, strong differences in survival were observed between specific pairings (P < 0.001), ranging from a 47.5% survival for pairing AA to complete mortality for pairings AE and EE (the survival curve of each pairing is provided in Figure S2). Alates from colony A had the highest survival rate, with 74 out of the 202 surviving alates originating from this colony (Figure 2a). Parings including an alate from A showed good survival overall (low hazard ratio), with the best survival observed for the inbred AA combination (Figure 2b). Notably, the opposite was also observed, with alates from colony E having the highest mortality rate. Consequently, pairings including an alate from this colony had low survival, with the lowest survival observed for the inbred pairing EE (Figures 2a&b). Overall, these results suggest that inbreeding has no effect on colony survival in the first several days after pairing; rather, the survival of the incipient colonies is strongly influenced by the colony of origin of the constituent partners.
The six colonies varied in their microbial loads, with colonies A, D and F exhibiting few CFUs (0.36, 0.39 and 0.58 for A, D and F, respectively; Figure 2d). In comparison, colonies B and C (and E to a lesser extent) displayed higher levels of microbial load with 14.86, 12.28 and 4.03 CFU, respectively. Interestingly, the susceptibility of a pairing was associated with the microbial load of the constituent colonies, both at the maximum microbial load (P = 0.0009) and the cumulative microbial load level (P = 0.0002; Figure 3a&b; Figure S3). The better fit of logarithmic regressions in both analyses suggests that hazard ratios only slightly increase after a certain threshold of microbial load (Table S1). In outbred pairings that included an alate from colonies B or C, the failure of the incipient colonies mostly resulted from the death of the alate from those colonies (Figure 2c), consistent with their elevated levels of microbial loads (the daily number and origin of dead alates are provided in Figure S2). In contrast, the opposite was found for outbred pairings including an alate from colonies A or D (low microbial loads), with the death of the partner originating from a different colony observed in most cases (Figure 2c,d and S2). Finally, the relationship between the relatedness of the partners and the hazard ratio of the colony pairing was not significant (P = 0.666), confirming the lack of effect of inbreeding on colony survival during the first 14 days after pairing (Figure 3c).
Bacterial communities were only slightly different between alates from different colonies on the PCoA (Figure 4a); weighted UniFrac values did not separate individuals from different colonies, while unweighted distances only moderately did (Figure 4a,b). Consequently, this results in similar levels of weighted bacterial differentiation observed within colonies and between different colonies (P = 0.733; Figure S4), and a lower, but non-significant, level unweighted differentiation within colonies than between colonies (P = 0.381). Fungal communities were also only moderately different between alates from different colonies (Figure 4a). The level of differentiation between nestmate and non-nestmate alates was significantly lower for weighted values (P = 0.045), but similar for unweighted values (P = 0.677; Figures 4a,b and S4). Overall, these results suggest that different colonies exhibit only slightly different bacterial and fungal communities. Consequently, only unweighted fungal dissimilarity between partners is marginally-significantly associated with an increase in hazard ratio of their pairing (P = 0.092; Figure 3g). However, the hazard ratio of a pairing was not associated with the weighted fungal similarity (P = 0.261), nor with the levels of either weighted or unweighted bacterial differences between partners (weighted:P = 0.478; unweighted: P = 0.862).