"Heart" cells may mask the effects of agrochemicals in bumblebees

The hepato-nephrocitic system can combat toxic substances depending on the concentration and length of exposure, researchers show.

By Elton Alisson

Agência FAPESP – The indiscriminate use of agrochemicals and disposal of toxic trace elements, including small amounts of heavy metals, in the soil, air, rivers and lakes are ranked among the factors responsible for the decline in bee populations and the disappearance of several bee species currently observed in different parts of the world.

The real effects of these chemical substances on insects have yet to be fully understood, however. No significant changes were detected by studies conducted in recent years in Brazil and other countries to find out whether exposing bees to varying concentrations of certain agrochemicals affects their mortality and survival, as well as their behavior, brain and other internal organs.

“When alterations are observed in mortality and behavior or in specific internal organs that may be affected by a given agrochemical, it’s not necessarily the case that the product is causing these effects in bees,” said Fábio Camargo Abdalla, a professor at the Biology Department of the Federal University of São Carlos (UFSCar), São Paulo State, Brazil, in an interview given to Agência FAPESP.

Abdalla and Caio Eduardo da Costa Domingues, a master’s student in UFSCar Sorocaba’s Graduate Program in Biotechnology & Environmental Monitoring with a scholarship from FAPESP, found that bumblebees (genus Bombus) have an integrated cellular system that is capable of “compensating” for the effects of toxicants. The insects are able to combat or “mask” the effects of these chemicals up to certain concentrations and exposure times.

The discovery, resulting from the research project “Effect of cadmium and Roundup® on internal organs of Bombus morio and Bombus atratus (Hymenoptera: Bombini)”, supported by FAPESP, was reported in an article published in the journal PloS One.

The findings will be presented at the next Latin American conference of the Society of Environmental Toxicology and Chemistry (SETAC), scheduled to take place in September in Buenos Aires, Argentina.

“Bees have this integrated cellular system, which we call the hepato-nephrocitic system, that was found to offset the effects of exposure to a certain xenobiotic,” Abdalla said. Xenobiotics are synthetic chemicals such as agrochemicals and trace metals found in living organisms and are not naturally occurring in the environment.

The hepato-nephrocitic system, identified by microscopy in bumblebees, Abdalla explained, comprises cells of the fat body, the functions of which correspond to those of the liver in humans, as well as pericardial cells and hemocytes (immune system cells).

All these groups of cells and tissues, layered in a myogenic contractile region around the dorsal vessel, which is the bee’s equivalent of a heart – a tubular sac in the abdomen that pumps the blood into the head –, function in a coordinated manner as a filter for the bee’s blood, which is called hemolymph.

When bumblebees are exposed to xenobiotics, the fat body cells are the first to be activated and represent an initial barrier against chemical aggression.

If the fat body cells fail to stop an “attack” by a chemical substance and are damaged or destroyed, the pericardial cells are activated.

However, the immune cell response occurs throughout the “combat” process. This is evidenced by cell morphology and blood cell counts during the entire period of exposure to agrochemicals or trace metals.

Toxic substances neutralized by the pericardial cells are released back into the hemolymph and can be filtered by the Malpighi tube, the insect’s excretory organ, which is equivalent to a mammal’s large intestine.

Bees’ immune cells participate throughout the process, Abdalla explained.

“The functions of this association of cells, together with the Malpighi tube, are analogous to those of the kidneys and liver in humans,” he said. “They represent the bee’s first line of defense to compensate for the harmful effects of exposure to synthetic chemicals.”

Possible biomarker

To determine the limit for compensation of the effects of toxicants by the hepato-nephrocitic system, the researchers performed experiments in which they exposed Bombus morio bumblebees to doses of cadmium considered environmentally safe in class 1 and 2 waters by Brazil’s National Environmental Council (CONAMA), as well as sublethal doses of thiamethoxam, the most widely used agrochemical in Brazil, and glyphosate for variable periods of time.

The bees’ cellular response was analyzed by counting cells from the hepato-nephrocitic system in their hemolymph. The analysis showed that exposure for two days to cadmium at 1 part per billion diluted in 2 milliliters of water caused fat body cell death and intense pericardial cell activity. This led to collapse of the system and destruction of the dorsal vessel.

“Our research shows that this also occurs in other species of bees aside from Bombus morio such as Bombus atratus and Apis mellifera, as well as the Xylocopa suspecta carpenter bee, which shares the same niche as Bombus but is solitary, whereas Bombus is social,” Abdalla said.

“This means that the hepato-nephrocitic system can be used as a morphological biomarker to analyze environmental stress levels in bees.”

Mere activation of the cells in this system by bees upon exposed to a certain type of xenobiotic, Abdalla explained, is itself an indicator of the harmful effects of the chemical, which forces the bees to divert metabolic energy that could be used for other purposes such as foraging.

“Colonies must be damaged if this energy deviation effect is extrapolated to all species of forager bees exposed in the wild,” he said.

According to Abdalla, analysis of the hepato-nephrocitic system can be used to predict quite accurately which organs may be affected by a given toxic agent, based on the cell types that are most damaged in the system, which performs several functions associated with other organs.

In addition to detoxification and filtering, Abdalla explained, the cells of the hepato-nephrocitic system are involved in ovarian development, in formation and maintenance of the cuticle, which is part of the bee’s exoskeleton (cuticulogenesis), and in the production of hormones that regulate brain glands.

“This system could be used as a checking point,” he said. “The classic methods used to study the effects of neonicotinoid insecticides, which are extremely harmful to the cells of a bee’s nervous system, may fail to detect morphological changes in the brain, not to mention changes in mortality and survival rates. However, it is possible to check whether this detox and filtering system is being activated.”

In the future, the researchers plan to use gas chromatography and mass spectrometry to analyze the fat body and pericardial cells that make up the hepato-nephrocitic system to investigate the dynamics of xenobiotic metabolization by bees.

Like the liver in humans, the bee’s metabolic system breaks down agrochemicals, for example, into smaller molecules called metabolites.

In certain cases these metabolites are far more powerful and cause more harm to the insect than the original agrochemical molecule, according to Abdalla.

“This is the case with thiamethoxam, which we’ve studied intensely in our laboratory,” he said. “When this agrochemical is ingested, its toxic potential can increase by a factor of 300.”

The article “Hepato-nephrocitic system: a novel model of biomarkers for analysis of the ecology of stress in environmental biomonitoring” (doi: 10.1371/journal.pone.0132349) by Abdalla & Domingues can be read in the journal PLoS One at www.plosone.org/article/related/info:doi/10.1371/journal.pone.0132349.