**1. Introduction**

Mozambique (Figure 1) is a country located in southeastern Africa (10◦30 –26◦52 S and 40◦50 –30◦31 E) covering a total land area of 800,000 km2. It is bathed by the Indian Ocean in the east and makes borders with Tanzania in the north; Malawi and Zambia in the northwest; Zimbabwe in the west and Swaziland and South Africa in the southwest.

**Figure 1.** Map of Mozambique. Red points or lines indicate the sites where Microcystin (MC) or MC producers were detected in Mozambique, and in near sites or in the shared rivers with Mozambique, Green points indicate the water sources or water treatment centers.

According to IV populational census carried out in 2017, this country has 29.67 million habitants distributed in 11 provinces [1]. The climate of Mozambique varies from subtropical climates (north and center) to dry arid (south) [2]. Like many African countries, Mozambique is highly vulnerable to climate variability and extreme weather events (droughts, floods, and tropical cyclones) [3]. Droughts are the most frequent natural disaster that have a negative impact on the population that reside in these rural areas [4]. The location of Mozambique in the coastal area makes it vulnerable to floods since many transnational river basins end [2]. Unfortunately, only 50% of the population has access to "safe drinking water". Urban areas are the most favored, with 80%, while rural and most of the population have only 35% coverage and consume untreated water daily from rivers, lakes, and small puddles that form after or during the rain [5–7], putting at risk public health.

Eutrophication of freshwater resources may lead to the occurrence of cyanobacterial blooms and the presence of cyanotoxins, being microcystins (MC), the most common toxins worldwide [8]. The presence of MC in untreated drinking water is a major threat to public health because this potent cyanotoxin causes hepatotoxicity in humans. Thus, this review evaluates the incidence of MC and its producers in drinking water bodies of Mozambique, based on reported and available data and the estimated human illness case numbers and associated economic damage caused by Microcystin both overall globally, and in Mozambique—Africa. Recommendations for routine control and monitoring of MC will also be done since this hepatotoxin is not included in water control tests data in Mozambique.

#### **2. Microcystin-Producing Species and Toxicology**

#### *2.1. Microcystin-Producing Species*

MCs are secondary metabolites produced by cyanobacteria species that occur naturally (but it can be increased severely by human activities) in freshwater environments. The most reported cyanobacteria species, which produce MCs are listed in Table 1 and include species of the families *Microcystaceae, Nostocaceae, Microcoleaceae, Oscillatoriaceae, Pseudanabaenaceae* (Table 1). The occurrence and development of a particular genus and species of cyanobacteria and cyanotoxins production worldwide seem to be conditioned to water chemistry and climate conditions [8]. In a temperate climate, *Microcystis* and *Anabaena* blooms occur widely while *Cylindrospermopsis* develops in tropical regions [9]. There are toxic and non-toxic cyanobacteria of the same species, which may be found together [8,10,11]. Toxic cyanobacteria can produce several toxins with different toxicity making it uncertain to assess the overall toxicity of bloom due to the variations of toxins concentration spatially and seasonally [12]. To distinguish toxic and non-toxic cyanobacteria species is very complicated, and consequently, the methods used are also complex. It implicates that the prevention of cyanobacteria bloom development is a suitable way to control toxic blooms [13,14].


**Table 1.** Microcystin-producing species detected in freshwater bodies.

The factors that promote the MC synthesis are not yet clearly understood, however, the optimal growth of MC-producing species and toxicity seem influenced by light intensity, nutrients, and temperature, among other factors. For example, the higher toxicity of *M. aeruginosa* extracts was verified in extreme pH values [39,40], and heavy metals such as Zinc and Iron did not influence the *M. aeruginosa* toxicity [41]. The content of nitrogen and phosphorus influenced the toxicity of *M. aeruginosa* extracts. Low nitrogen content reduces the *M. aeruginosa* toxicity, while low phosphorous increased the toxicity in the natural population [42,43] and reduced in lab experiments [16,21,44,45]. Another lab conclusion was the correlation of colony size and content of toxic cyclic heptapeptide of the non-axenic strain of *M. viridis* and axenic *M. viridis* was also verified [20,46,47]. In general, the optimal temperature for which MC-producing species produce MC ranged from 20 to 25 ◦C [21,40,48,49]. This range of optimal temperature suggests that cyanobacteria blooms are most toxic during periods with warm weather and in areas with warm climates [8].

#### *2.2. Toxicology*

Microcystins (Figure 2) are the largest diverse group of cyanobacterial toxins, and to date, more than 240 MCs analogs are known, and they vary structurally in terms of the degree of methylation, hydroxylation, epimerization, peptide sequence, and consequently in their toxic effects [50–52]. Chemically, MC is a group of monocyclic heptapeptides (numbered in Figure 2) containing both D- and L-amino acids plus N-methyldehydroalanine (Mdha) and a unique β-amino acid side-group, 3-amino-9-methoxy-2-6,8-trymethyl-10-phenyldeca-4,6-dienoic acid (Adda) and their analogs differ among them, at the two L-amino acids and on the methyl groups on D-erythro-β-methylaspartic acid (D-MeAsp) and Mdha with molecular weight varying from 900 to 1100 Daltons. MC-LR, MC-RR, and MC-YR are common MC variants, the letters L, R, and Y represent the aminoacids leucine, arginine, and tyrosine, which appear on the MC molecule in different combinations [50,53–58] being MC-LR the most studied. The biosynthesis of this group of cyanotoxin is regulated by non-ribosomal peptide synthetase and polyketide synthase domains, being *MCyS* the gene cluster, which has been sequenced and partially characterized in several cyanobacterial species of the family *Microcystaceae, Nostocaceae, Microcoleaceae, Oscillatoriaceae, Merismopediacea,* and *Pseudanabaenacea* [16–38,59,60]

**Figure 2.** General chemical structure of microcystins. The common MC variant is MC-LR when X and Y correspond to L-Leu and L-Arg.

The mechanism of MCs toxicity seems to be well understood. They bind to serine/threonine-specific protein phosphatases (PPs) such as PP1 and PP2A, inhibiting their activity [61–63]. Adda moiety (Figure 1) plays an important role in the MC toxicity group since its isomerization and/or oxidation reduces the toxicity [64,65]. The inhibition of PP1and PP2A as a result of MC acute exposure causes excessive protein phosphorylation, alterations in the cytoskeleton, loss of cell shape, and consequently destruction of liver cells leading to intrahepatic hemorrhage or hepatic insufficiency [58]. Oxidative stress increasing in cells and consequent apoptosis, which can cause tumor promotion, is another mechanism of MC toxicity [66–68].

#### **3. E**ff**ects of Microcystin in Humans, Symptoms, and Treatment**

Microcystin effects in humans depend on the time of exposure and concentration ingested [69], and the studies are based on epidemiologic data but the reported studies on laboratory animals. Human health problems are mostly caused by chronic exposure by consumption of contaminated water or food, dermal exposure, or inhalation [57]. MC human poisoning episodes were reported in different parts of the world after the consumption of contaminated water or during sport or recreational activities [59,70–72]. Some examples of MC human poisoning cases are described; America—in 1996, an episode of human intoxications by MC was reported in Brazil with more than 76 deaths of patients at two dialysis centers in Caruaru. The municipal water supplied to the dialysis centers was the source of MC [71,73–75]. In Argentina, a human poisoning caused by MC involving a young man after immersion in an intense bloom *Microcystis* sp lake during sport and recreational activities were recorded. Four hours after exposure, the patient showed nausea, abdominal pain, and fever, and 48.6 <sup>μ</sup>g·L−<sup>1</sup> of microcystin-LR was detected in the water samples [76]. Other cases were recorded in Uruguay (January 2015) involving a 20-month-old child and her family during recreational activities. These victims were admitted to the hospital with diarrhea, vomiting, fatigue, and jaundice and the analysis confirmed the presence of MC-LR (2.4 ng·g−<sup>1</sup> tissue) and [D-Leu1]MC-LR (75.4 ng·g−<sup>1</sup> tissue) explanted liver [77]. Africa—toxic cyanobacteria suspected intoxication cases were reported in

Zimbabwe involving children that were hospitalized in the Hospital of Harare with gastroenteritis symptoms [78]. In Europe—121 people presented abdominal pain, nausea, vomiting, diarrhea, fever, headaches, and muscle pain after consumption of untreated water from the River Kavlingean in Sweden. In this case a bloom of MC—producing such as *Planktothrix agardhii* and *Microcystis* spp. was observed in the river [79]. The most affected human organ is the liver [57]. However, in vivo and in vitro studies indicated that the kidney and colon are also affected [80–85]. The symptoms generally reported in humans due to the MC intoxication include gastroenteritis and related diseases, allergic and irritation reactions, liver diseases, tumors, and primary liver cancer and colorectal cancers, and massive hepatic hemorrhage. MC human poisoning treatment is very complicated due to the rapid, irreversible, and severe liver damage [86], however, gastric lavage [87], administration of monoclonal antibodies against MC-LR [88], immunosupressant Cyclosporine A, antibiotic rifampin [89], and membrane-active antioxidant vitamin E, taken as a dietary supplement [90] are recommended.
