Stingless bee native to Brazil cultivates fungus to survive

Larvae of Scaptotrigona depilis depend on a microorganism to complete their development cycle, Brazilian researchers have discovered.

By Elton Alisson

Agência FAPESP – Researchers have discovered that Scaptotrigona depilis, a species of stingless bee native to Brazil (common name mandaguari), cultivates a fungus to survive. The larvae of these bees feed on the fungus, which is similar to a species of fungus used by humans for centuries as a food preservative, especially in Asia.

The discovery, described in an article published on October 22 in the online edition of the journal Current Biology, resulted from PhD research performed by Cristiano Menezes with a scholarship from FAPESP.

“This is the first time that symbiosis has been observed between a social bee species and a cultivated fungus,” said Menezes, a researcher at the Eastern Amazon branch of the Brazilian Agricultural Research Corporation (EMBRAPA), based in Belém, Pará, and first author of the paper.

“We know some species of ants and termites have symbiotic relationships with fungi they cultivate in their own nests. These microorganisms provide their hosts with nutrients and protect them against pathogens. But this is the first time such a relationship has been found in bees,” Menezes told Agência FAPESP.

The study was part of the Thematic Project “Biodiversity and sustainable use of pollinators, with emphasis on Meliponini bees”, coordinated by Vera Lucia Imperatriz-Fonseca, a researcher at the Vale Sustainable Development Technology Institute (ITVDS) and Full Professor at the University of São Paulo’s Bioscience Institute (IB-USP).

The researchers found that the newborn larvae of this stingless bee species feed on filaments of a fungus in the genus Monascus (Ascomycotina) placed in their nest.

Without this microorganism, which produces several secondary metabolites with anti-microbial, anti-tumoral and immune defense properties, few larvae survive, according to the authors of the study.

“We don’t yet know exactly what this fungus does for the larvae. The most plausible hypothesis is that it helps protect their food from pathogens, given its use in China and other parts of Asia as a food colorant and preservative,” Menezes said.

International media coverage of the research includes a story published by Newsweek.

Transmission to next generation

According to Menezes, the fungus grows out of the brood cell walls, which are made of cerumen, a mixture of wax produced by worker bees and plant resins.

When worker bees finish building the nest, the brood cells are filled with a semi-liquid food mass regurgitated by nurse bees. The queen then lays an egg on the top of the food in each cell, which is sealed by workers and reopened only when the adult bee emerges. The fungus starts proliferating when the egg is about to hatch, about three days after being laid. It grows out of the cerumen, spreads over the surface of the semi-liquid food and is devoured by the larva, disappearing completely by the sixth day after hatching.

“Our recordings of the behavior of three-day-old larvae showed that they cut the fungal mycelia with their mandibles and ingested them,” Menezes said.

The fungus is passed on to the next generation when the bees found a new colony by swarming, a process that, in stingless bees, uniquely involves taking building materials and stored food from the mother nest.

Once the larvae have left their brood cells, workers start scraping the cerumen to reuse it as material for a new nest. In addition, during swarming they transport cerumen from the mother colony to build brood cells in the daughter colony. Fungal mycelia do not grow until the brood cells are ready and the reused cerumen comes into contact with the larval food deposited in them by nurses.

“We also don’t yet know whether spores are involved, or parts of the mycelia themselves lie dormant in the cerumen and are conveyed from one brood cell to another,” Menezes said.

He observed the same reliance on fungi to complete the birth cycle in other stingless bee species in the genus Scaptotrigona, as well as species of Tetragona, Melipona and Frieseomelitta.

“This kind of mutualism between bees and microorganisms appears to be much more frequent than has been thought and should make us more concerned about the use of fungicides in agriculture,” Menezes said.

Studies performed in recent years in the US and Europe have shown that fungicides are among the pesticides most often found in bee pollen.

“The worry is that fungicides could be killing microorganisms beneficial to bees, like the fungus identified in this study,” he said. “If these chemicals are present in bee pollen, they inevitably reach brood cells.”

Accidental discovery

Menezes discovered the symbiosis between S. depilis and Monascus by accident. During his PhD research in entomology at the University of São Paulo in Ribeirão Preto, conducted with a scholarship from FAPESP, he developed a method for mass producing queens of S. depilis in the lab in response to demand from farmers for more colonies of the species, which pollinates several crops.

To produce queens, Menezes enriched the diet of female larvae because what determines whether a female larva of this species will become a worker or a queen is diet in the larval phase (read more at: http://agencia.fapesp.br/18536).

He raised female larvae in artificial brood cells kept in a humid chamber with large amounts of enriched food. After a few days, he observed a white fungus beginning to grow out of control, and then the larvae died.

“At first I thought the fungus was causing a disease in the bees,” he said. “I tried to exterminate it with chemicals and remove it mechanically, but nothing worked.”

Sometime later, however, he noticed the fungus growing less rampantly in natural brood cells. “It seemed as though something in the bees’ natural environment was keeping the fungus under control,” he said.

When he tried raising the female larvae under less humid conditions, he found that the fungus grew intensely for a few days and then disappeared.

“After that, I obtained a survival rate of more than 90%. I suspected that the female larvae were feeding on the fungus and depended on it to survive,” Menezes explained.

To test this hypothesis, the researchers performed experiments in which two groups of larvae were reared in the lab, one on sterilized larval food alone and the other on sterile food supplemented with fungal mycelia.

The larvae with fungus-enriched food displayed a 76% survival rate, while for those raised in the same conditions but without the fungus the survival rate was only 8%.

“This points to a very strong dependency of the bees on the fungus,” Menezes said.

At the same time, the fungus benefits from being cultivated in the nests of this stingless bee species because its reproduction is guaranteed for generations.

“Apparently, the bees are the main beneficiaries of this symbiosis,” he concluded, “but the fungus also depends on them to reproduce.”

The article “A Brazilian Social Bee Must Cultivate Fungus to Survive” (doi: 10.1016/j.cub.2015.09.028) by Menezes et al. can be read in Current Biology at www.cell.com/current-biology/abstract/S0960-9822(15)01108-2.