Data from: Eggshells as hosts of bacterial communities: an experimental test of the antimicrobial egg coloration hypothesis

Oviparous animals have evolved multiple defenses to prevent microbes from penetrating their eggs and causing embryo mortality. In birds, egg constituents such as lysozyme and antibodies defend against microbial infestation, but eggshell pigments might also impact survival of bacteria. If so, microbes could exert an important selective pressure on the evolution of eggshell coloration. In a previous lab experiment, eggshell protoporphyrin caused drastic mortality in cultures of Gram positive, but not Gram negative, bacteria when exposed to light. Here, we test this ‘photodynamic antimicrobial hypothesis’ in a field experiment. In a paired experimental design we placed sanitized brown, protoporphyrin-rich chicken eggs alongside white eggs that lack protoporphyrin. We deployed eggs for 48 hrs without incubation, as can occur between laying and incubation, when microbial infection risk is highest. Eggs were placed on the open ground exposed to sunlight and in dark underground storm-petrel burrows. We predicted that the proportion of Gram positive bacteria on brown eggs should be lower when exposed to sunlight than when kept in the dark, but we expected no such difference for white eggs. Although our data revealed variation in bacterial community composition, the proportion of Gram positive bacteria on eggshells did not vary by egg colour, and there was no interaction between egg colour and location. Instead, Gram positive bacteria were proportionally more common on eggs on the ground than eggs in burrows. Overall, our experiment did not support the photodynamic antimicrobial hypothesis. The diverse range of avian egg colours is generated by just two pigments, but over 10 hypotheses have been proposed for the evolution of eggshell colour. If our results are generalizable, eggshell protoporphyrin might not play a substantial role in defending eggs against microbes, which narrows the field of candidate hypotheses for the evolution of avian eggshell coloration.

卵生动物已演化出多种防御手段，以阻止微生物侵入卵内并造成胚胎死亡。对于鸟类而言，溶菌酶、抗体等卵内成分可抵御微生物侵染，而蛋壳色素或许也会对细菌的存活产生影响。若确有此效应，微生物便可能对蛋壳颜色的演化施加重要的选择压力。此前的实验室实验显示，蛋壳中的原卟啉（protoporphyrin）在光照条件下可使革兰氏阳性（Gram positive）细菌的培养物出现大幅死亡，而对革兰氏阴性（Gram negative）细菌则无此影响。本研究通过野外实验对这一“光动力抗菌假说（photodynamic antimicrobial hypothesis）”进行验证。我们采用配对实验设计，将消毒后的富含原卟啉的棕色鸡蛋，与不含原卟啉的白色鸡蛋一同放置。实验中，我们将卵放置48小时且不进行孵化，以模拟产卵至孵化前的时段——该时段的微生物感染风险最高。卵被分别置于暴露于阳光的开阔地面，以及黑暗的地下风暴海燕（storm-petrel）洞穴中。我们预测，暴露于阳光下的棕色鸡蛋表面的革兰氏阳性细菌占比，应低于置于黑暗环境中的个体，但预期白色鸡蛋不会出现此类差异。尽管我们的数据显示细菌群落组成存在差异，但蛋壳表面的革兰氏阳性细菌占比并未随蛋壳颜色发生变化，且蛋壳颜色与放置位置之间不存在交互效应。相反，地面放置的卵上的革兰氏阳性细菌占比，要高于洞穴内的卵。总体而言，本实验并未支持光动力抗菌假说。鸟类多样的卵色仅由两种色素生成，但目前已有超过10种假说被提出以解释蛋壳颜色的演化。若本研究结果具备普适性，那么蛋壳原卟啉或许并未在抵御微生物的过程中发挥显著作用，这一结论将缩小关于鸟类蛋壳颜色演化的候选假说范围。