Keywords

1 Introduction

Planet Earth has been facing an immense ecological crisis for decades. The environmental decay has both economic and environmental effects on its inhabitants. One of the majorly affected ecosystems that are crucial for the proper functioning of the planet is the aquatic ecosystem. Climate change, pollution, farming and other anthropogenic factors have caused the rapid worsening of the water quality over the recent years [7]. This trend became a threat to many organisms inhabiting the aquatic environments but also to humans who are almost entirely dependant on the natural fresh water supplies. An efficient and robust monitoring and global sharing of the water quality data is the most effective method of preventing the ecosystem collapse. Aquatic monitoring has been practiced for a long time with the use of various sensors and sample taking. While these methods are most accurate and reliable in terms of the predictability of the outcome, they are time and money consuming [22]. Classical sensors can be costly and require frequent calibration and maintenance, therefore a continuous monitoring is close to impossible. An urgent need for more efficient methods of aquatic monitoring has arisen. The door to long-term environmental monitoring opens with the use of robots. Robotic entities are able to reach not easily accessible habitats and remain there for long periods of time continuously gathering large amounts of data. One way to achieve a long-term energy-efficient monitoring is combining the electronic parts with living organisms to create so-called “biohybrids” [17]. Initiatives surrounding this topic have been carried out in the recent years [10, 18, 20]. Biohybrids for long-term aquatic monitoring were implemented in a European Union funded project “Subcultron” [20]. Through the use of MFCs as a natural power source, bacterial activity was used to gather data on the environment by tracking their electricity patterns. By using the swarm strategy, the robots were able to collect large amounts of data from many places simultaneously, providing a better overview of the investigated place. This project created three types of robots: aMussel, aFish, aPads, each with their own purpose in the robotic swarm.

This innovative project succeeded on performing various unsupervised marine monitoring missions. MFC was used as a power source for the aMussels which were embedded in the sediment. The microbial activity enabled the aMussel to operate autonomously and also provided an insight into the benthic processes by monitoring the power output [20]. The concept of using a living organism as a sensor in biohybrid entities was extended with the project Robocoenosis [15,16,17]. Project Robocoenosis introduced here, aims to take a new perspective on this methodology and expand it with the use of multiple locally found lifeforms, such as mussels, Daphnia spp., Hydra spp., various larvae etc. [15]. Here, is where Robocoenosis uses the extreme complexity of the aquatic environments to its advantage.

The intricacy of the aquatic environments makes investigating one parameter at a time, which results in a very precise data, quite inefficient long-term. Ultimately, it might not accurately portray the overall condition of the water body. Lifeforms are in constant contact with the surrounding environment and respond to the overall of factors and changes occurring within it. Moreover, lifeforms do not require frequent maintenance as under normal conditions, they sustain themselves in their natural habitats. Lifeforms employed as live biosensors can be mussels, Daphnia, benthic community structure and many others [15]. Here, core features of the project Robocoenosis and its various applications are presented [15,16,17].

1.1 Robocoenosis Biohybrid Entities

Project Robocoenosis aims to develop novel methodology of utilising lifeforms as live sensors in place of the classical sensors with the use of biohybrids [17]. By combining the technical and electrical parts with the live sensors provided by organisms a complex biohybrid entity is able to hack into the ecosystem and obtain reliable data. The entities developed within this project will allow the scientific community to obtain long-term data on the status of the selected lake. This will be achieved by the low-power electronics and self-charging module with the use of Microbial Fuel Cells (MFC).

Autonomy. The aim for these prototypes is for them to be self-powering. To ensure the power autonomy the robot is supplied with MFCs. MFCs are one of many types of fuel cells [1]. They rely on the anaerobic breathing processes of the bacteria present in anoxic sediments. They use locally available organic matter to produce low-current electricity which can be accumulated and used by the biohybrid entity [1].

Low-Power. Due to the low currents obtained by the MFC, this method will only be used for short periods of time. To overcome the power shortage, the internal electrical parts (such as a single board computer “Raspberry Pi”) are turned off or put into a sleep mode between the data collection periods. Low-power micro-controllers and other micro devices will be used.

Biodegradability. On top of the features mentioned above, the biohybrid entities designed by Robocoenosis are meant to be semi-biodegradable. This is accomplished by constructing two segments: 1) non-biodegradable core containing all the necessary electronic parts and 2) biodegradable platforms hosting organisms and the lifeforms themselves. After the monitoring mission is terminated, the non-biodegradable core will return to the surface and will be collected by the researchers while the biodegradable parts will remain in the environment.

2 Methods

2.1 Fields of Work

Various Austrian lakes have been chosen as the field of work. Each of them represents a vastly different habitat to make the biohybrid testing relevant to the a broader variety of surroundings. Firstly, Lake Millstatt (N\(46^\circ 47'\) E\(13^\circ 34^\prime \)) was chosen as a classic example of an oligotrophic lake, that is, a lake with a low amount of nutrients and clear, well-oxygenated waters. Glacial lakes, such as Lake Millstatt, are the ones created due to the glacial activity, whether being carved out by the glacier or supplied by the melt-water [4].

Lake Neusiedl (N\(47^\circ 56'\) E\(16^\circ 50^\prime \)) represents a completely different habitat. It is highly eutrophic, i.e. contains a high content of nutrients and algae. It is also likely to be met with anoxic conditions near the bottom sediments [19]. While being the largest lake in Austria surface-wise, it is also extremely shallow, being less than 2 m deep at the deepest point. These characteristics make this body of water extremely vulnerable to changing temperature and nutrient inflow.

Another example of a glacial lake that will be used for the field experiments is Lake Hallstatt (N\(47^\circ 34'\) E\(13^\circ 39^\prime \)) located in Upper Austria. The nutrient content is slightly higher than in Lake Millstatt and thus, this lake is classified as oligo-mesotrophic meaning that its water quality parameters fall in between oligo- and eutrophic conditions and leaning towards the former [8, 9]. All of the lakes mentioned above are protected and highly sensitive areas. Biohybrid entities allow for an extensive and long-term environmental monitoring of these water bodies with minimal human involvement.

2.2 Lifeforms

Daphnia. When using a lifeform’s physiological or behavioural responses as an indicator, it is essential to choose a well-documented species in order to obtain a reliable result [15]. One of the invertebrates most widely used as bioindicator is Daphnia sp. Because of its sensitivity to various chemicals and changes in the environment, it has become a popular choice for toxicological studies, behavioural and pharmaceutical research [6]. In case of a stressor present, certain reactions, such as swimming speed, vertical distribution or heart rate, can be used to identify its presence and intensity [14]. Daphnia’s swimming behaviour is a good indicator of its health as affected individuals may display decreased swimming speed, increased sinking and other behaviours. By closely monitoring their swimming patterns there is a high chance of detecting a stressor in the environment. In Robocoenosis, this is achieved by constructing a flow-through chamber in which Daphnia can swim freely in the enclosed layer of water (Fig. 1). Thanks to an appropriately-sized mesh build into the setup, animals can still easily feed on the algae naturally occurring in the water. Daphnia specimens were sampled from the respective lakes to avoid introducing alien species to the environment. A camera piece controlled by a single board computer (Raspberry Pi) included in the setup is facing the chamber. The setup is lowered with suspension cables the bottom of the respective lakes at various depths.

Fig. 1.
figure 1

Early biohybrid prototype during field test in the Lake Neusiedl. The prototype includes A) a chamber containing swimming Daphnia, B) a Raspberry Pi camera module and C) a Raspberry pi contained within the plexiglass tube.

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Zebra Mussel. Another species of focus is the zebra mussel Dreissena polymorpha. It is a relatively small species with the size ranging between 2–2.5 cm, and most commonly lives in large colonies. It thrives in well-oxygenated waters and thanks to its resilience it is now widely spread across the world and was labeled as highly invasive by the Invasive Species Compendium [3, 5]. As zebra mussel feeds and breaths through filtrating the surrounding waters it responds quickly to any changes in the environment or the presence of toxic substances with certain stress behaviours [11]. A sudden appearance of an undesirable factor such as low oxygen, temperature spike or a toxic substance will result in an instant closing or fluttering movements of the valves. When the observed specimen presents any of those behaviours, it is a sign of a stressor present in the environment. In Robocoenosis, data from this behaviour is harvested by combining visual observation and automated movement tracking (Fig. 2). A mussel is mounted to the setup with a tag on the top valve to augment the valve movements. The mussel is recorded at a set interval to observe the movements and any potential signs of stress.

Fig. 2.
figure 2

A: Zebra mussel Dreissena polymorpha with a red tag mounted on the top valve in order to facilitate the movement detection. B: Preliminary results of the object tracking done in Lake Millstatt.

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3 Discussion

The project Robocoenosis introduces a new approach to environmental monitoring. Robocoenosis aims to develop novel methodologies to using living organisms as natural biosensors. This opens the door to an efficient long-term collection of large amounts of data with minimal human impact on the environment. The outcomes of this project will find many applications in environmental management and biological research, as well as exploring the interactions between lifeforms and the environment. The use of biohybrids will allow to monitor bodies of water that would otherwise be inaccessible or where sampling surveys are strictly limited. It will also provide a new methodology for ethological and ecological research. Through the robotic setup it is possible to observe the animals in their natural habitat without the usual disruptions of in-person observation, such as gas bubbles or movements while handling or maintenance.

Over the years, various lifeforms were used as biosensors. Daphnia has been extensively used, for example, by pharmaceutical companies for testing the effects on cardioactive drugs on the heart rate [21]. Daphnia’s light responses were also successfully employed to detect small quantities of toxic chemicals in the water [13]. A disruption in the overall swimming behaviour including clustering, swimming speed and distribution, was treated as an early-warning signal by Daphnia Toximeter II [12]. When the behaviour of enough individuals deviates from normal, an alarm is triggered. With these laboratory-based uses of Daphnia’s sensitivity, it is of merit to investigate its uses in the field. Robocoenosis will move the toxicological studies directly into the water body of interest decreasing significantly the time of detection and sample processing period [2].

As mentioned before, this was successfully achieved by the project Subcultron based in Adria region [20]. With the use of a robotic swarm equipped with various sensors, the researchers were able to gather data not only in a continuous manner but also cover large areas at the same time [20]. This initiative will be taken a step further with the project Robocoenosis.

With regards to ethics, project Robocoenosis will use exclusively invertebrates which do not fall into the scope of animal rights laws. Regardless of this, minimally invasive techniques will be investigated.

Here, major aims of the project Robocoenosis have been presented. Its aim is designing a novel methodology to investigating the aquatic environments while focusing on long-term and autonomous setups. This will be done with the use of biohybrids, combining robotic parts with living organisms.