The different olfactory worlds of female and male moths

A team of researchers from the Max Planck Institute for Chemical Ecology, together with colleagues from Martin Luther University in Halle-Wittenberg, Germany, studied the olfaction of female silk moths. Using electrophysiological methods, they discovered that the antenna, specialized in males to detect female pheromones, is particularly sensitive to the smell of silkworm droppings in females.

The components of this odor were found to be a deterrent to mated females, likely allowing them to avoid competition for their own offspring when laying eggs. The responsible sensory neurons are located in hair-like structures called sensilla.

In males, pheromone detection takes place in a long type of these sensilla, while neurons in the long sensilla of females detect the odor of larval feces. The odor of mulberry, the silkworm’s only host plant, is on the other hand detected by the sensory neurons of female butterflies in the sensilla of medium length.

The study is published in the journal Proceedings of the Royal Society B: Biological Sciences.

The world smells different to females than it does to males.

In humans, the sense of smell is developed equally in men and women, although women have slightly more olfactory neurons and therefore a slightly more sensitive nose. But overall, they perceive the same smells. Male butterflies, on the other hand, live in a completely different olfactory world than their female counterparts. For example, the antennae of male silk moths – their “noses” – are highly specialized to detect female sex pheromones, whereas females cannot even smell their own pheromones.

There are thousands of sensilla on the antennae, hair-like structures, which can be divided into morphologically and functionally distinct groups. The most common sensilla in males are long and contain two sensory neurons. One specializes in detecting bombykol, the female sex pheromone, while the other responds to bombykal, a component of the pheromone of other butterfly species. While bombykol is very attractive to male butterflies, bombykal is a deterrent.

“Since females cannot smell their own pheromone, it has long been thought that their long sensilla also have a very specific function found only in females. After mating, the female’s only task is to find a suitable plant on which to lay it. It was therefore suggested that the long sensilla of females are specialized to detect the attractive scent of mulberry trees. We wanted to test this hypothesis,” explains Sonja Bisch-Knaden, who leads a project group at the Department of Evolutionary Neuroethology at the Max Planck Institute for Chemical Ecology.

The long sensilla of female silk moths recognizes silkworm droppings

Electrophysiological methods, such as measuring the activity of each sensilla (monosensilla recording), were crucial to the study results. The scientists not only tested many different individual odors, but also mixtures of natural odors, such as those found in mulberry leaves, caterpillar droppings, the body odor of moths or meconium, a liquid that butterflies secrete when they hatch.

All these odors, which play an ecological role in the silk moth’s environment, had been collected. The research team was also able to match the expression of olfactory receptors to the corresponding type of sensilla.

“We were surprised to find that the neurons in the long sensilla of female butterflies were not specialized to detect the odor of the host plant, as expected, but that one of the two neurons in the long sensillum is very sensitive to odors such as isovaleric acid and benzaldehyde. Detection of the odor of the mulberry leaf itself is carried out by medium-length sensilla neurons,” explains Bisch-Knaden.

Isovaleric acid and benzaldehyde are odorous components of silkworm droppings. Using a simple Y-maze test with an input arm that splits into two side arms through which an odor or clean air (control) is introduced, the research team was able to elicit behavior in otherwise immobile females that expressed attraction or aversion. Major differences emerged when comparing virgin and mated females.

The researchers showed that odors associated with caterpillar droppings did not trigger a specific reaction in virgin females, but had a deterrent effect on mated females. Presumably, the smell of feces helps females avoid mulberry trees, which are already teeming with silkworms when they lay their eggs.

In search of the male silk moth pheromone

The pheromone of female silk moths, bombykol, was chemically characterized as early as 1959 – the first insect pheromone ever created. So far, scientists have not been able to identify a male counterpart. The current study provides clues, but no answers to the question of a male pheromone.

“The second neuron in the long sensilla of females is highly specific for (+)-linalool, an odor previously identified as a component of the male pheromone in other butterfly species. However, no linalool could be found in “Body odor of male silk moths, and (+)-linalool alone had neither attractive nor repellent effects on female moths in behavioral experiments,” says Bisch-Knaden.

Peculiarities of silk moth odor perception

By studying the molecular basis of odor detection in female butterflies, researchers noticed a particularity in the spatial organization of olfactory receptors. There are two families of olfactory receptors, the evolutionarily older ionotropic receptors (IRs), which detect primarily acids, and the odorant receptors (ORs), which detect a wide range of chemically diverse compounds. Based on studies in the model fly Drosophila melanogaster, it has long been thought that neurons expressing IRs or ORs generally occur in different types of sensilla and that IRs never appear in long sensilla.

In the silk moth, however, an IR co-receptor for acid detection and the obligate OR co-receptor are both found in the same neurons located in the long sensilla. This co-expression of IRs and ORs increases the chemical receptivity of sensory neurons. Odors detected by both types of receptors are processed and transmitted together, which could be advantageous for the unambiguous detection of ecologically important odor mixtures.

“It is amazing that research into insect olfaction continues to produce surprising results. Our study shows that it is important to study more than just one model,” says Bill Hansson, head of the Department of Evolutionary Neuroethology.

The researchers also discovered this co-expression of the two types of receptors in the long sensilla of male butterflies, which is why they speculate that acid sensing could also play an important ecological role in males. Further investigation will now clarify this.