The volatile alkylpyrazines methyl- and methoxypyrazines (MPs) present in the reflex bleeds of coccinellid beetles such as the harlequin ladybird beetle Harmonia axyridis are important semiochemicals that function in antipredatory defense behavior. Pyrazines have also been coadapted from a primarily defensive role into pheromones that function in intraspecific communication, attraction, and aggregation behavior. However, the biosynthesis of MPs in ladybird beetles is poorly understood. Here, we tested the hypothesis that MPs could be produced by microbial symbionts in H. axyridis, which generates four different MPs. The evaluation of tissue-specific MP production showed that MP concentrations were highest in the gut tissue and hemolymph of the beetles rather than the fat body tissue as the presumed site of MP biosynthesis. Furthermore, manipulation of gut microbiota by antibiotic-containing diets resulted in a lower MP content in adult beetles. The analysis of the bacterial community of the digestive tract revealed the presence of bacteria of the genera Serratia and Lactococcus which are reportedly able to produce MPs. In line with the known diet-dependent production of MP in H. axyridis, we determined that the presence or relative abundance of some of the potential MP producers (Enterococcus and Staphylococcus) is also diet-dependent. We hypothesize a potential role of the microbiota in MP production in H. axyridis as a possible example for outsourcing the synthesis of ecologically important semiochemicals to its gut bacteria.
Keywords: Harmonia axyridis; Lactococcus lactis; chemical defense; gut microbiota; methoxypyrazines (MPs); symbiosis.