Occurrence of Harmful Cyanobacteria in Drinking Water from a Severely Drought-Impacted Semi-arid Region

Abstract

Harmful cyanobacterial blooms have become increasingly common in freshwater ecosystems in recent decades, mainly due to eutrophication and climate change. Water becomes unreliable for human consumption. Here, we report a comprehensive study carried out to investigate the water quality of several Campina Grande reservoirs. Our approach included metagenomics, microbial abundance quantification, ELISA test for three cyanotoxins (microcystin, nodularins, and cylindrospermopsin), and in vivo ecotoxicological tests with zebrafish embryos. Cytometry analysis showed high cyanobacterial abundance, while metagenomics identified an average of 10.6% of cyanobacterial sequences, and demonstrated the presence of Microcystis, Cylindrospermopsis, and toxin coding genes in all ponds. Zebrafish embryos reared with pond water had high mortality and diverse malformations. Among the ponds analyzed, Araçagi showed the highest lethality (an average of 62.9 ± 0.8%), followed by Boqueirão (lethality average of 62.5 ± 0.8%). Here, we demonstrate that water from ponds undergoing extremely drought conditions have an abundance of potentially harmful cyanobacteria and their toxins. Our findings are consistent with a scenario in which polluted drinking water poses a great risk to human health.

Keywords: cyanotoxins; eutrophication; metagenomics; public healthy; zebrafish.

FIGURE 1

General overview of the sampling. (A) Position of the sampling locations. The study was carried out in ponds located in Paraíba State (Brazil), which have been affected by a persistent dry period. Filled circles denote pond sites. Water was sampled approximately 0.5 m below the water surface twice during September and October 2016. (B) Principal component analysis of the ponds based on chemical and biological variables showing differential features of each pond. Variables (arrows) and sampling sites (Araçagi, Boqueirǎo, Saulo Maia, Galante, and Mazagǎo). Control is represented by the square. The sampling ponds occupied different ordination space composed by PC1 and PC2.

FIGURE 2

Abundance of sequences related to Cyanobacteria and cyanotoxins. (A) The major cyanobacterial genera found in the metagenomes of Araçagi, Boqueirão, and Saulo Maia ponds. (B,C) Abundance and distribution of genes involved in the biosynthesis of cyanobacterial toxins found in the metagenomes Values correspond to the relative abundance for the total identified clusters of orthologous groups of proteins sequences (COGs), corresponding to COG 1020 sequences. Microcystin (mcy) and cyanopeptolin (mcn) synthetase gene clusters were found, as well Anabaenopeptilide/-peptins gene cluster (apd) (B) and non-ribosomal peptide synthase genes and other genes potentially involved in the biosynthetic pathways for toxins production (C). Araçagi, Boqueirão, and Saulo Maia major ponds presented mcy, mcn, apd, other, and non-ribosomal peptide synthase genes. COG proteins assigned to Anabaenopeptilide/-peptins gene cluster were all annotated as Anabaena sp. 90, whereas Microcystis aeruginosa was related to cyanopeptolin synthetase gene cluster for all ponds (exception Planktothrix spp. for McnC in Boqueirão 1). M. aeruginosa was also related to microcystin synthetase gene cluster in all ponds. For non-ribosomal peptide synthetase, the major sequences were related to M. aeruginosa, Nodularia spumigena, Aphanizomenon flos-aquae, Nostoc spp., Microcystis panniformis, Planktothrix spp., Anabaena spp., Fischerella spp., Scytonema spp., and Chroococcidiopsis thermalis. Control, bottled mineral water.

FIGURE 3

Environmental water ponds induce different developmental defects in zebrafish. Teratogenic effects were observed at 96 and 120 hour post-fertilization (hpf) (N = 60 embryos for each treatment). (A,B) Developmental delays; malformations of the head, mouth, jaw, spine, and tail; and heart edema were observed in zebrafish treated with Araçagi water samples. (C,D) Developmental delays and malformations of the spine, tail, tail curving, heart edema, and deterioration were observed in zebrafish treated with Boqueirão water samples. (E,F) Normal development observed in zebrafish controls. (G) Developmental observations rate in zebrafish. Unaffected and teratogenic rates observed in embryos treated with environmental water samples. Most embryos from Saulo Maia were unaffected, while most embryos from Araçagi and Boqueirão presented lethality, followed by different malformations. (H) Developmental observation rate of teratogenic effects in Araçagi and Boqueirão.