Why Are Brine Shrimp Used in Experiments? The Complete Guide for 2024

Artemia, Stress, and Society

Response to stressful situation maximizes fitness advantages of animals in nature, survival, behavior, and reproduction being the obvious outcome. Artemia for example is an example of stress response of an animal low in the tree of life that, however, exhibits similarities to humans that are up in the biological hierarchy making evident that evolution builds complex solutions from simpler ones, and such solutions heavily depend on the complexity of the environment. The Artemia environment is relatively simple in terms of the number of known stressors or stresses affecting it, though it is clear the environment is multidimensional. Instead, the human physical and cultural environment has evolved faster than adaptation to them, and the consequence is a maladaptive response or disease (adaptation syndrome; see McEwen, 2007). Alike Artemia, the brain plays a key role in coordinating the behavioral and physiological response to stressors. Another striking phenomenon is the ability of Artemia females to recognize and select peers as a way to maintain ecological adaptation, which is somewhat found in humans as well.

Finally, understanding the mechanisms of adaptation to stressful environments in Artemia has some indirect benefits to human societies because of the role it plays in the aquaculture of marine fish and crustaceans (Dhont and Sorgeloos, 2002). Larval stages of some of these species cannot utilize pelleted first feed but need a live diet. Artemia larvae (which can be nutritionally enhanced) provide not only basic nutritional requirements but also enzymes and other valuable dietary elements as well forming an attractive prey for predatory fish larvae. Artemia production is a highly profitable industry. In the developed world cysts are harvested from sites such as the Great Salt Lake in Utah and after canning and vacuum packing are sold in quantity worldwide. In developing nations such as Vietnam and Thailand, cheaper cysts are produced in artisanal saltworks.

As a major macro-faunal inhabitant of salty lakes, the brine shrimp Artemia provides a unique example of how to develop a strategy in dealing with critical conditions needed for survival. Being adapted to the harsh conditions that salty lakes impose on survival and reproduction of individuals, but also on populations and species, constitutes a complex evolutionary response that integrates different levels of biological organization. Such a highly integrated response has evolved from more simple solutions in a process that requires the constant availability of innovations (through mutation and other genomic events) and the filtering process of natural selection. The relatively high diversity of prokaryotic life in salty lakes is an example of simpler though extremely successful solutions, but also the co-existence of sexual species and parthenogenesis in Artemia, shows the importance of variation in reproductive modes for evolution to proceed. Parthenogenesis has been successful under certain environmental conditions, some of which remain in certain areas or lakes as to allow this reproductive mode to persist. The long time co-existence of parthenogens and the sexual species from which they originated, is a phenomenon that some biologists (at least of those working on salty lakes) find it difficult to accept considering the widespread adoption of sexual reproduction by most taxa.

The following aspects of salty lakes and the extremophile Artemia, the two relevant actors in our evolutionary play, are highlighted in this article:

• Salty lakes offer a quite restrictive range of ecological conditions, salinity being often recognized as a critical stressor. However, additional stressors such as specific ionic conditions also favor the differentiation of locally adapted populations (ecological isolation) that thrive in chloride, carbonate, or sulfate-rich lakes. A better characterization of salty lakes should provide further details on how adaptation proceeds in Artemia.
• The ability of females to “perceive” forthcoming environmental difficulties is amazing though it is an example of the signaling process species have evolved through a highly interactive and dynamic process. This is yet a field to be explored.
• The ability of females to switch offspring quality, i.e., to produce cysts or nauplii, is a good example of the importance of the above (2). Cysts and nauplii are equipped with a differential suite of adaptations to cope with the environment immediately ahead, in the case of nauplii (often a stable one), or in relatively uncertain immediate environments or that to be faced in the years to come (Rode et al., 2011).
• Cysts are arguably the most resistant of all animal life history forms to environmental stress, whilst motile stages (nauplii, larval stages, adults) are the best osmoregulators in the animal kingdom as stated by Clegg and Trotman (2002). Cysts are gene banks that store a genetic memory of historical population conditions, but also are survival vehicles. They aid in the dispersal of Artemia, but also are reservoirs of genetic variability (Gajardo and Beardmore, 1989), the fuel for evolutionary change and resilience.
• There are few genetically divergent Artemia species restricted to specific regions (regional endemism), but highly divergent populations adapted to the specific ecological conditions of salty lakes (see 1). Hence, it seems the strategy of the species to persist is to distribute the gene pool in different baskets (salty lakes) that are well adapted to more specific conditions.
• To avoid loss of these adaptations by genetic flux (a role attributed to water birds and wind), the island-like nature (geographical isolation), and the local ecological pressures in particular prevent immigrants from surviving and, therefore, prevent or limit cross-breeding.

Salty Lakes, only for Salt-Lovers

Hypersaline lakes are distributed in all continents even in the Antarctic, mostly in tropical and sub-tropical areas where solar radiation is high enough to favor high evaporation rates which are required to maintain high salt concentration. But striking exceptions exist, such as the very high altitudes of Chile and Tibet. There are currently about 500 sites were Artemia has been reported (Van Stappen, 2002) but it is certain that many more exist around the world. Their recognition as extreme ecosystems becomes evident from their low eukaryotic biodiversity, to the point that, in a few cases, Artemia is the only macroscopic representative. Hence, Artemia is rightly referred to as a model animal extremophile or “halophile” (salt-lover; Wharton, 2007), whilst the relatively high prokaryotic biodiversity observed in salty lakes (DasSarma and Arora, 2001; Demergasso et al., 2004) indicates different evolutionary capabilities or strategies in these organisms. Salty lakes with Artemia are, therefore, considered good biodiversity laboratories due to their simple web structure (Gajardo et al., 2006). Nevertheless, Artemia tolerates very well the very large environmental variations that exist in many salty lakes, achieving very large population sizes. The range of environmental variation is due in some degree to latitude and the associated climatic conditions. Some salty lakes are permanent, where Artemia flourish year-round, while others are seasonal and dry-out in a predictable or unpredictable manner (Lenz and Browne, 1991). Some locations are coastal or thalassohaline (NaCl major salt) while others are far inland, athalassohaline (rich in anions other than chloride), such as those that are found at 4,500 m above sea level in the Tibetan Plateau. This area has been recently highlighted as the “third pole” because it has the third largest reserve of ice on earth after the Arctic and Antarctic, and it is alarming that the ice is melting quickly (Qiu, 2008). Closer to the stratosphere than any other Artemia site, this is a special location subject to high UV radiation and peculiarly cold for its latitude. In the Tibet area around 352 saline lakes exist (Van Stappen, 2002). By contrast at the other extreme of the globe, the Atacama Desert in Chile is one of the driest areas in the world, and Artemia is found there at about 2,500 m altitude in a unique environmental setting (Gajardo and Beardmore, 1993; Demergasso et al., 2004). In contrast to the melting down of ice in the Tibet by indirect human causes like climate change, salty lakes in the Atacama Desert are increasingly perturbed by water drainage associated with mining activities (deposits of NaCl and lithium salts). Two other quite unusual Artemia locations are found in Chilean Patagonia (Amarga and Cisnes lagoons; Gajardo et al., 1999, 2002; Clegg and Gajardo, 2009; Beristain et al., 2010) that experience severe weather conditions, rather unusual for Artemia standards (cold, rainy, and extremely windy). To complete this very brief account of Artemia environmental variability, there are also man-made salt ponds or salinas, where salt is produced by evaporation of seawater, and where Artemia can exist in certain salty ponds that approach the precipitation point of NaCl, 340 g L−1(Clegg and Trotman, 2002). However, in order for Artemia reproduction to occur the salinity must be well below that, although still very high.

Characteristic examples of natural lakes are found in Asia in areas such as China, Tibet, Iran, and Kazakhstan. All these contain four of the seven sexual Artemia species currently known, making evident both the peculiarities of salty lakes and the ecological divergence or specificity of Artemia species. The Mediterranean area, where Artemia probably originated, contains the fifth sexual species (Abreu-Grobois and Beardmore, 1982; Abreu-Grobois, 1987; Gajardo et al., 2002; Van Stappen, 2002; Baxevanis et al., 2006), and the other two species are found in Lake Grassmere in New Zealand, Chaplin Lake in Canada, the Great Salt Lake in Utah, USA, and the already mentioned Chilean sites. These are just examples – there are many other cases.

Although salinity seems to be the major driver of Artemia distribution, other critical factors are particularly severe in specific areas, as evidenced above. Most of them have synergistic effects, difficult to evaluate in nature, but amenable to study under laboratory conditions (Lenz and Browne, 1991). Salinity, ionic composition and temperature are the critical factors highlighted by Van Stappen (2002), whilst Hebert et al. (2002) additionally considers UV radiation as being of utmost evolutionary importance, since it increases mutation rates, the novelty factor, and/or the raw material, on which natural selection acts. Accordingly, Hebert et al. (2002) observed higher rates of DNA sequence divergence in the halophilic Anostraca from Australia and North America compared to their fresh-water relatives (Daphniids).

For the purpose of this article we group these critical factors as follows:

• (i)Reversible stressors, when reaching the upper or lower limit tolerated by Artemia (temperature and salinity).
• (ii)Mutagenic stressors, affecting DNA replication fidelity (ionic composition and UV light, but salt concentration plays a role as well). On the positive side they could generate variability (new functional variants), whilst their negative effects are filtered out by selection.

Salinity ranges widely (1–mM–5 M) (Hebert et al., 2002) because of latitudinal variation and other considerations, and so Artemia can be found with other planktonic animals at low-medium salinities. The optimum is estimated under laboratory conditions at 60 g L−1 in experiments where several fitness parameters were compared (Lenz and Browne, 1991), the maximum being close to NaCl saturations in solar ponds (340 g L−1), whilst the lower limit depends on the upper salinity tolerance of fish predating on Artemia, in extreme cases being as high as 100–130 g L−1 as reported by Van Stappen (2002). As it will be shown later in Section “Artemia: A Survival Machine” Artemia has evolved efficient solutions to these challenges, from molecules to fractal physiological processes (see also Figure 2). A major impact of high salinity is osmotic stress, desiccation, lowoxygen tension, increased metabolic rate to cope with the high energy demand required to maintain the osmoregulatory systemat full capacity, and alteration of DNA-protein relationship that lowers DNA replication fidelity (Hebert et al., 2002).

The temperature box ranges widely (5–40°C), the lower limit being consequences of the extreme habitats already discussed such as those in the Tibetan Plateau, Atacama Desert, and Patagonia, in northern and southern Chile, respectively. The upper limit is often seen in man-made salinas where Artemia thrives in shallow ponds. The optimal temperature for at least some species of Artemia has been established at about 25°C (Lenz and Browne, 1991; Van Stappen, 2002), but extremely high temperatures can overlap the effects of extreme salinity that drastically reduce the size of Artemia populations. Extremely low temperatures, including freezing, are overcome by the production of encysted embryos (see below). Depending on the prevailing anions, salty lakes are classified as chloride, sulfate, or carbonate-rich, including the possibility of combinations of two or even three major anions. Variation in ionic composition is therefore wide and likely the highest among metazoans (Cole and Brown, 1967; Van Stappen, 2002). Ultraviolet light is normally high in salty lakes as these are mainly located in areas of high solar radiation, but it is particularly high in those at high altitude due to the reduction in stratospheric ozone. UV radiation reduces metabolic activity and induces the formation of dimers of DNA more efficiently at 254 nm. UV-B (280–320 nm) produces irreversible damage and death, and the effect is greater in naupliar stages than adult (lower LD50; Dattilo et al., 2005). The ability to repair such mutations explains, in part, differential survival observed between individuals and populations.

In addition to the molecular-cellular and physiological impact of these stressors on individuals, there can be population/species effects such as ecological divergence (Schuler and Conte, 2009). This has been recognized as a speciation mode in Artemia by Abreu-Grobois (1987), Mono Lake in California, USA, a carbonate-rich lake described as an evident example of ecological isolation (Browne and Bowen, 1991).

Edmonton students blasting their brine shrimp experiment to the International Space Station

FAQ

Why are brine shrimp good for experiments?

… animals because they allow students/teachers to quickly demonstrate the effects that such nutrients may have on the health, growth, or survival of organisms

What is the point of brine shrimp?

Brine shrimp are a valuable food source to migratory birds that congregate in and around the Great Salt Lake.Mar 20, 2023

What is the main purpose of the brine shrimp assay?

Now-a-days brine shrimp (Artemia salina, fairy shrimp or sea monkeys) lethality assay is commonly used to check the cytotoxic effect of bioactive chemicals.Jun 5, 2017

What are common uses of brine shrimp in aquaculture and scientific research?

Brine shrimp have the ability to produce dormant eggs, known as cysts. This has led to the extensive use of brine shrimp in aquaculture. The cysts may be stored for long periods and hatched on demand to provide a convenient form of live feed for larval fish and crustaceans.