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Occurrence
Anatoxin-a is produced by multiple genera of cyanobacteria that are found in fresh water, brackish, and marine environments globally. Cyanobacterial blooms (sometimes referred to as cyanoHABs) are increasing in frequency globally. This is caused by increasing water temperatures and eutrophication because of nutrient runoff from sources like agricultural fertilizer, livestock waste, and septic tank overflows. These blooms increase the amount of cyanotoxins in the surrounding water, threatening the health of both aquatic and terrestrial organisms. Some species of cyanobacteria that produce anatoxin-a are Aphanizomenon, Anabaena, Raphidiopsis, Oscillatoria, Planktothrix, and Cylindospermum.

Deserts
Cyanobacteria are key primary producers, and in desert environments, occur in puddles after seasonal rains on ephemeral river beds and supertidal salt flats; however desert cyanobacterial growth is far less studied. While crust analysis has led to the detection of microcystins, production of anatoxin-a and anatoxin-a(S) isn't clear due to acetylcholine esterase inhibitors and further studies are needed.

Recreation and Exposure
Anatoxin-a is an alkaloid neurotoxin. Most poisonings in humans are tied to recreational activates in lakes or rivers when swimming in water, inhaling water spray, or swallowing water where these blooms are present. Human exposure can also occur in dietary substances. However, there could be severe public health impacts should these blooms occur in drinking water supplies or agricultural irrigation. Some response cases are due to allergic responses, while others are cases of neurotoxicity or respiratory distress. Cyanobacterial toxins can block ion-channels, create neuromuscular blockades, anti-acetylcholinesterase activity, and anti-phosphatase activity, and protein synthesis inhibition. These can cause liver damage, tumor growth, and gastrointestinal issues.

Recreational Exposure
The World Health Organization (WHO) has created a maximum level of cyanotoxin allowed in drinking water and has created guidelines for recreational waters. Cyanobacteria is a growing concern for human health and there are very few counties with regulations in place to protect humans.

The Great Lakes
There have been many neurotoxin producing blooms in cyanobacteria species, however most studies are conducted of microcystins, saxitoxin and neosaxitoxin. Anatoxin-a has been detected in low concentrations in the western basin of Lake Erie and embayments of Lake Ontario.

Lake Dianchi
PCR and qPCR were used to determine the presence and amount of anatoxin-a producing bacteria. Aphanizomenon and Anabaena are two common cyanobactera found in Lake Dianchi. Aphanizomenon was found to be the main producer of anatoxin-a and are most abundant through algal blooms during the spring and summer months. Anatoxin-a producing Aphanizomenon were found in the lowest concentrations near the north end of the lake and increased southward.

Other Cases of Toxicity
In 2003, along the La Loue River shorelines in eastern France there were rapid deaths of dogs due to the biofilm that built up on the surface of the water and stones that contained many many benthic species of filamentous cyanobacteria that produced anatoxin-a. Anatoxin-a was discovered using high-performance liquid chromatography (HPLC) with UV detection by electrospray ionization-Quadrupole-Time-Of-Flight mass spectrometer and mass spectrometry. Phormidium favosum was the cyanobacteria that produced anatoxin-a which was the cause of the toxicity in these dogs.

A young, male Golden Retriever presented with acute onset paraparesis 40 minutes prior. After swimming in a man-made pond in Reno, Nevada, the dog continued to smell of pond water 5.5 hours later and was non-ambulatory with generalized ataxia that was most noticed in its pelvic limbs. At presentation the dog was anxious with mild ptyalism, tachycariac, normothermic, and eupneic. When excited, the dog experienced muscle tremors and eventually progressed to latera recumbency. After examining the area for potential toxins, the owner noted the dog ingested 2.5 cups of algae that were removed from the pond. After 2 hours the dog suffered from mental depression, tensor rigidity, and absent menace before the dog was ventilated. Although the dog regained consciousness, the dog lacked menace and pupillary light reflexes.

In-vitro Exposure
This study discusses the effects of in vitro exposure to anatoxin-a. Anatoxin-a is good for studying nicotinic receptors as it is a neurotoxin that acts as a nicotinic acetylcholine receptor agonist. Anatoxin-a caused dopamine to be released in a concentration-dependent way, but perfusion of nicotinic antagonists (mecamylamine and abungarotoxin) prevented striatal dopamine release. Anatoxin-a can be used as a nicotinic antagonist by in vivo microdialysis. When stimulated by anatoxin-a, dopamine release is partially inhibited when receptor antagonists (metillycaconitine or a-bungarotoxin) are perfused.

This study examines the effect of the synergistic relationship between anatoxin-a and algal toxin microcystin-LR may have on in virto embryotoxicity in mice and toads. The mice babies (pups) had no significant effects on viability or weight when born. These pups were monitored after birth to follow their neurological developments, but no neurotoxicity was observed. At all concentrations of toxin, there were no deaths and no post-birth neurotoxin developmental delays. In toads, dose-dependant transient narcosis, edema, and loss of equilibrium were observed. In high doses, there was a high mortality rate a few days after exposure.

Mice
In mice the LD50 of anatoxin-a was found to be 200 mg*kgK1 body weight via intraperitoneal injection. When administered by a single-dose gavage (orally), anatoxin-a is shown to have and LD50 of 5 μg.g−1 of body weight. The no observed adverse effect level (NOAEL) by the oral route over a 28 day period was found to be 0.12 mg kg71 bodyweight per day, although the authors note that the actual NOAEL may be 3.0 mg kg71 bodyweight per day since dosing was done with hydrochloride (HCl).

Fish
In this experiment, medaka fish were exposed to anatoxin-a orally to best simulate how they would naturally come into contact with the neurotoxin through biofilms on lake surfaces. Individuals that had doses of up to 6.67μg.g-1 survived and were able to recover without any symptoms of toxicosis fairly quickly when placed back into freshwater. On the otherhand, individuals with anatoxin-a doses of up to 20μg.g-1 rapidly presented neurotoxic effects which included decreases of ofopercular movement, abnormal swimming, and global musculature rigidity. Some individuals eventually expressed sporadic breathing that stopped after 15 minutes, but all fish exposed to 20μg.g-1 showed no sighed of recovery. In medaka fish, the LD100 is 20μg.g-1, the no observable adverse effect level (NOAEL) was 6.67μg.g-1 and the LD50 was calculated to be 11.5μg.g-1.

Dogs
The LD50 in dogs is not known as cases of anatoxin poising bring on rapid and acute effects that quickly kill the dog, making it difficult to study. Once exposed, dogs suffer from acute neurological issues after about 5-10 minutes and usually die within 1 hour.

Research Gaps
Compared to other cyanotoxins like microcystins, anatoxin-a is understudied and because of this there is a lack of knowledge on fresh water systems and toxic events that can occur.

Occurrence and Biogeography
Many studies of anatoxin-a occur in wealthier counties as a response to visible cyanobacterial growth or public health issues in heavily populated areas; however, anatoxin-a toxicity can still occur in areas where there is no evident cyanobacterial growth. In addiction, the cost of analyzing and collecting samples of anatoxin-a and saxitoxin could exclude less affluent countries. Most experiments south of the equator have only occurred in Australia and New Zealand which excludes many freshwater environments. Furthermore, more research in less populated areas would be beneficial to learn more about the biogeography and ecosystem effects.

Environmental Fate and Degradation
There are very few studies that examine the fate of neurotoxins in benthic or terrestrial soil.

Exposure Routes
There are many studies about exposure through drinking water, recreational activities, or consuming fish or shellfish that have been contaminated, very little research on inhalation (that could be caused from storms, rainfalls, or insects). Little understanding of secondary exposure in freshwater such as food preparation. Understanding secondary effects will lead to more understanding of effects on various trophic levels.

Triggers of Neurotoxin Production and Release
Understanding what causes neurotoxin production and release will lead to better predictors of when and where exposures will occur. The effects of salinity, temperature, sunlight, pH, and nutrient availability are not often discussed in studies or data are not related water quality. Furthermore, collecting primary data and doing toxin analysis' will determine toxin production and release.

Food web Effects
Freshwater studies of food web effects are primarily focused on microcystin and many food web effects in marine environments have been conducted on saxitoxin. There is very little research on food webs or natural communities to examine the sublethal effects of neurotoxins like anatoxin-a.

Studies of Cyanotoxin Mixtures
Many toxins can be produced in a bloom from a variety of toxin-producing strains of cyanobacteria co-existing at once. Laboratory studies would be beneficial to observe the effects of multiple toxins as it is not know if these neurotoxins have a synergistic or antagonistic effects with other cyanotoxins.

Sublethal Effects of Neurotoxins
It is known that there are mild to severe health effects from anatoxin-a and saxitoxin exposure that most commonly occur from recreational actives in water (e.g. swimming or boating). Most studies agree that the lethal does for anatoxin-a is between 0.20-0.25 mg/kg; however, these levels are not likely to occur in one sitting and acute toxicity is unlikely. Therefore more studies should be conducted that address the sublethal health and ecosystem effects of chronic freshwater neurotoxin exposure from recreation actives in water.