Science of the Total Environment 756 (2021) 143697

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Science of the Total Environment

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Immediate and long-term impacts of one of the worst mining tailing dam
failure worldwide (Bento Rodrigues, Minas Gerais, Brazil)
Cristiane dos Santos Vergilio a,⁎,1, Diego Lacerda b,1, Tatiana da Silva Souza a, Braulio Cherene Vaz de Oliveira b,
Vinicius Sartori Fioresi a, Victor Ventura de Souza a, Giovana da Rocha Rodrigues a,
Marjore Kinaip de Araujo Moreira Barbosa a, Echily Sartori b, Thiago Pessanha Rangel b,
Diogo Quitete Ribeiro de Almeida b, Marcelo Gomes de Almeida b,
Fabiano Thompson c, Carlos Eduardo de Rezende b,⁎⁎
a
Laboratório de Ecotoxicologia, Departamento de Biologia, Centro de Ciências Exatas Naturais e da Saúde, Universidade Federal do Espírito Santo, Campus Alegre. Alto Universitário, S/N,
Guararema, Alegre, Espírito Santo CEP: 29.500-000, Brazil
b
Laboratório de Ciências Ambientais, Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense Darcy Ribeiro, Avenida Alberto Lamego, 2000, Parque Califórnia, Campos
dos Goytacazes, Rio de Janeiro CEP: 28.013-602, Brazil
c
Laboratório de Microbiologia, Centro de Ciências da Saúde, Instituto de Biologia, Universidade Federal do Rio de Janeiro, Ilha do Fundão, Anexo ao bloco A, Rio de Janeiro CEP: 219.449-70, Brazil

H I G H L I G H T S G R A P H I C A L A B S T R A C T

• The Fundão dam failure is one of the

largest environmental disasters in the
world.
• Immediately the tailings were mainly
transported as suspended particulate
matter.
• After 6 months, with the deposition, the
metal levels increased in the sediment.
• Water and sediment induced negative
biological and toxicogenetic effects.
• This joint of abiotic and biotic indicators
can direct other monitoring programs.

a r t i c l e i n f o a b s t r a c t

Article history: The rupture of Fundão Dam is considered one of the largest environmental disasters in Brazilian history and
Received 7 July 2020 one of the largest in the world involving tailings dams. The present study analyzed the changes in metal
Received in revised form 21 October 2020 concentrations in the dissolved, suspended particulate matter (SPM) and sediment in the period just
Accepted 2 November 2020
after (15 days) and six months after the dam rupture, together with the biological and cytogenotoxic ef-
Available online 23 November 2020
fects, from the collapse site until the Doce River mouth in the Atlantic Ocean. After the dam rupture, the tail-
Editor: Daniel A. Wunderlin ings were mainly transported as SPM. After six months, with the deposition, there was a decrease in metal
concentrations in dissolved and SPM and increased levels were observed in the sediment. Cr, Ni, Cd and Hg
Keywords: levels in sediment were higher than the threshold effects level (TEL/NOAA), especially six months after the
Heavy metals dam rupture. The water induced immediate negative biological effects at different levels of the trophic
Ecotoxicology chain, together with Al, Fe, Mn and Zn accumulation in fish muscle. Both water and sediment also showed

⁎ Correspondence to: C. S. Vergilio, Laboratório de Ecotoxicologia, Departamento de Biologia, Centro de Ciências Exatas, Naturais e da Saúde, Universidade Federal do Espírito Santo -
Campus Alegre, Alto Universitário, S/N, Guararema, Alegre, Espirito Santo 29.500-000, Brazil.
⁎⁎ Correspondence to: C. E. de Rezende, Laboratório de Ciências Ambientais, Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense Darcy Ribeiro, Avenida
Alberto Lamego, 2000, Parque Califórnia, Campos dos Goytacazes, Rio de Janeiro 28013-602, Brazil.
E-mail addresses: cristiane.vergilio@ufes.br (C. dos Santos Vergilio), crezende@uenf.br (C.E. de Rezende).
1
C. S. Vergilio and D. Lacerda have the same contribution as first authors.

https://doi.org/10.1016/j.scitotenv.2020.143697
0048-9697/© 2020 Elsevier B.V. All rights reserved.
C. dos Santos Vergilio, D. Lacerda, T. da Silva Souza et al.
Iron ore cytotoxic, genotoxic and mutagenic effects. These data demonstrate the importance of long-term monitor-
Sediment ing with abiotic and biotic parameters to clarify the impacts of mining tailings and can help to direct future
Suspended particulate matter monitoring programs.
1. Introduction
Mining activity produces intense changes in the landscape to re-
cover valuable minerals. The process involves extraction of large
amounts of rock and soil, with intensive use of water, resulting in re-
lease of effluents such as tailings (Schoenberger, 2016). The material hy-
draulically retained by tailings dams is usually composed of fine
particulate matter with greater weight (ICOLD, 2001). The volume and
chemical characteristics of the tailings released by dam failure (particu-
larly with potentially toxic metals) damage the whole affected area,
including soils, rivers, plants, associated biota and local human popula-
tions (Burritt and Christ, 2018).
On November 2015, the rupture of Fundão Dam, located in south-
eastern Brazil, initially released at least 34 million m3 of iron ore mining
residue, which flowed for more than 650 km through the Gualaxo do
Norte, Carmo and Doce rivers until reach the Atlantic Ocean (IBAMA,
2015; Carmo et al., 2017) (Supplementary Video). Efforts were made
to retain the remaining 16 million m3 in the tailings pond, but a consid-
erable part also reached the sea. The volume of material released
and resulting damage made this event the largest environmental disas-
ter in Brazilian history, and one of the largest in the world involving
tailings dams (Aires et al., 2018; Almeida et al., 2018; Burritt and
Christ, 2018).
The volume of tailings led to an immediate increase in water turbid-
ity, contamination of riverside soils, loss of native vegetation, including
remnants of Atlantic Forest (a hotspot of biodiversity) and other places
legally classified as “permanent protection areas” (Aires et al., 2018;
Omachi et al., 2018). The damages were severe to water, sediment
and associated fauna of the river courses, as well as the estuarine,
coastal and marine environments (IBAMA, 2015; Carmo et al., 2017).
Besides environmental damages, the social impacts were also severe,
with the death of 19 people, pollution of the water supply, evacuation
of the riverside population, restrictions on fishing activities and other
water uses (such as irrigation), socioeconomic losses and sense of dan-
ger and helplessness among the affected population (IBAMA, 2015;
Carmo et al., 2017; Hatje et al., 2017). Nowadays, the population is
still fearful of the contamination risk due to the lack of transparency of
the mining company and public environmental agencies.
The flow of tailings released not only the characteristic elements
from the ore, predominantly Fe (45,200 ± 2850 mg·kg−1) and Mn
(433 ± 110 mg·kg−1), but also toxic metals as Cr (63.9 ±
15.1 mg·kg−1), Pb (20.2 ± 4.6 mg·kg−1) (Queiroz et al., 2018), Hg
(~75 ng·g−1), As (~719.07 ng·g−1) (Segura et al., 2016) and even the
rare earth metals La (~414.57 ng·g−1) and Sc (~609.30 ng·g−1)
(Segura et al., 2016). Despite efforts to quantify and monitor the impacts
of the disaster on inland and coastal ecosystems, many uncertainties re-
main more than four years after the dam rupture. The initial impact of
the tailings as source of metals was demonstrated, but the fluctuations
in metal levels with time have not been confirmed yet and the long-
term effects are not clearly described. This material was eventually de-
posited in riverside soils and river bottom sediments, which can be
absorbed by organisms of the local food chain.
Among the biological consequences of tailings release are changes in
microbial communities (Cordeiro et al., 2019), algae physiology (Costa
et al., 2019), benthic macrofauna (Oliveira Gomes et al., 2017) and larval
fish assemblages (Bonecker et al., 2019), beyond alteration in liver
morphology of the fish species (Weber et al., 2020). However, some of
these studies are restricted to marine or estuarine areas or only cover
2
Science of the Total Environment 756 (2021) 143697
© 2020 Elsevier B.V. All rights reserved.
restricted areas of the Doce River. Considering the magnitude of the
accident, there are still few studies reporting the immediate and long-
term impacts of the tailings release on biological organisms. Therefore,
there is still a need to evaluate the impacts along the affect area, includ-
ing evaluation of the metal accumulation in the biota, to enable infer-
ences about the continuing risks to the human population.
Toxicological tests using different indicator organisms are an impor-
tant tool for water quality monitoring. The use of species with distinct
position level can help to assess the impacts along the food chain.
Genetic toxicity bioassays are also recommended to monitor the quality
of impacted areas, because they shed light on the action of compounds
with cytotoxic, genotoxic and/or mutagenic properties, like heavy
metals. Nevertheless, only a few studies have focused on the biological
and cytogenotoxic potential the compartments of the Doce River after
the iron ore tailings release. Moreover, many of the studies were
restricted to one or two sampling sites (Gomes et al., 2019; Quadra
et al., 2019). Therefore, considering the dimensions of the disaster and
its potential impacts on exposed organisms and human populations, it
is important to investigate the biological and cytogenetic impact of the
tailings in the Doce River as a function of time and distance from
the dam.
The present study investigated the changes in metal concentrations
in the dissolved, suspended particulate matter and sediment in the
period just after (15 days) and six months after the rupture of the tail-
ings dam, together with the biological and cytogenetic effects of the
tailings, from the collapse site until the mouth of the Doce River at the
Atlantic Ocean. These data are important to understand the metal
dynamics along time and the long-term biological impacts of this
massive accident, to support actions to restore and protect the Doce
River watershed and similar impacted areas worldwide.
2. Material and methods
2.1. Study area and sampling
Fundão Dam belongs to the Germano mining complex, in the Bento
Rodrigues district located in the municipality of Mariana in Minas Gerais
State, southeastern Brazil. The dam was storing some 50 million m3 of
iron ore tailings, of which at least 34 million m3 was released directly
into the Doce River basin (Supplementary Video). On November 5,
2015, with the dam rupture, the tailings reached the Santarém Dam
just in downstream, then flowed through the Gualaxo do Norte River
until reaching the Carmo River and ultimately the Doce River (Supple-
mentary Video). Along the Doce River, the tailings were carried for
670 km until the Atlantic Ocean, in the municipality of Linhares in
Espírito Santo State (IBAMA, 2015).
The Doce River basin is an area of great social and environmental
importance for Brazil, between Minas Gerais and Espírito Santo states,
containing 228 municipalities, an estimated population of more than
three million inhabitants (ANA, 2020). The region has the largest ore
mining complex in Latin America, supporting the economy of several
municipalities. Besides mining, other uses of the Doce River include
supply of drinking water for humans and livestock, irrigation, and
fishing, among others (IBAMA, 2015).
The region has humid tropical climate with a rainy season from
October to March and a dry season from April to September (ANA,
2020). The headwaters of the Doce River's tributaries are at altitudes
above 1000 m. Along its course, mainly beyond the city of São Jose do
C. dos Santos Vergilio, D. Lacerda, T. da Silva Souza et al.
Goiabal, the Doce River flows at altitudes lower than 378 m (Supple-
mentary Material S1).
To evaluate the elements' distribution and biological effects of the
tailings along the river, water and sediment samples were collected
from near the rupture site (2.5 km) until the river mouth (666 km) in
two campaigns: (1) in the period immediately after the dam collapse
(November 23–27, 2015 – when the tailings first reached the sea) -
with 9 sampling sites (Ipatinga – 256 km, Governador Valadares –
365 km, Tumiritinga – 407 km, Resplendor – 475 km, Aimorés –
500 km, Mascarenhas – 523 km, Colatina – 556 km, Linhares – 622 km
and Regência – 666 km; and (2) six months after the dam rupture
(May 9–12, 2016) – with 13 sampling sites (Bento Rodrigues –
2.5 km, Gualaxo – 71 km, São José do Goiabal – 157 km, Bom Jesus do
Galho – 242 km, Ipatinga – 256 km, Governador Valadares – 365 km,
Tumiritinga – 407 km, Resplendor – 475 km, Aimorés – 500 km,
Mascarenhas – 523 km, Colatina – 556 km, Linhares – 622 km and
Regência – 666 km). Along the river until the Atlantic Ocean there are
four water dams: (1) between Gualaxo and São José do Goiabal, (2) be-
tween Ipatinga and Governador Valadares, and (3) and (4) between
Aimorés and Mascarenhas (Fig. 1).
At each sampling site, the physical-chemical parameters pH, electri-
cal conductivity, dissolved oxygen and turbidity were measured
(Table 1). One aliquot (500 mL) from the surface water filtered within
a day of sampling using pre-weighted glass fiber filters (0.70 μm) to ob-
tain the suspended particulate matter (SPM) through gravimetry for
metal analysis as well as organic matter composition (Table 1), followed
by water acidification until pH 2 and storage in polyethylene containers
at 4 °C for dissolved metal analysis. After filtration, the glass fiber filters
Fig. 1. Map of the sampling sites along the Doce River basin in southeastern Brazil. The sampling started close to the Fundão Dam rupture site and followed along the Gualaxo, Carmo and
Doce rivers until the mouth at the Atlantic Ocean. 1 - Bento Rodrigues – 2.5 km, 2 - Gualaxo – 71 km, 3 - São José do Goiabal – 157 km, 4 - Bom Jesus do Galho – 242 km, 5 - Ipatinga –
256 km, 6 - Governador Valadares – 365 km, 7 - Tumiritinga – 407 km, 8 - Resplendor – 475 km, 9 - Aimorés – 500 km, 10 - Mascarenhas – 523 km, 11 - Colatina – 556 km, 12 - Linhares –
622 km and 13 - Regência – 666 km).
3
Science of the Total Environment 756 (2021) 143697
that contained SPM were freeze-dried and stored at −20 °C for further
analysis. Another the surface water aliquot (15 L) was collected and
stored in polyethylene containers at 4 °C for biological and toxicogenetic
assays. The surface sediments were collected in plastic bags and kept
under refrigeration at −4 °C until analysis. One sediment portion was
fractionated (< 2 mm) by sieving, after a wet sample from the fraction
smaller than 2 mm was placed under agitation in an ultrasound bath
for 10 min, followed by the determination of the particle size distribu-
tion by laser diffraction (Shimadzu model SALD-3101) in sand, silt and
clay fractions (Table 1). Another sediment portion (<2 mm) was
homogenized with a mortar and pestle for subsequent metal
measurements.
2.2. Metal quantification in dissolved, SPM and sediment
Metal analysis in the dissolved fractions were performed in the acid-
ified water samples during the sampling campaigns. SPM filters and
sediment (0.5 g dry weight) were prepared according to the US EPA
(1996) 3052 method through acid digestion (HNO3 65%, HF 48% and
HCl 37%) in a microwave oven (Mars 5 Xpress CEM-Corporation). A
4% boric acid (H3BO3) solution (m/v) was added to complex the possible
HF residue. The extracts were filtered and the tubes were filled to 50 mL
with HNO3 0.5 M. Metal determination (Al, Ba, Cd, Ce, Cr, Cu, Er, Eu, Fe,
Gd, La, Lu, Mn, Nd, Ni, Pb, Pr, Sc, Sm, Tb, Ti, Y, Zn) was performed by in-
ductively coupled plasma-optical emission spectrometry (ICP-OES,
Varian - 720 ES), while Hg determinations were performed in a Hg
Quick Trace M-750 CETAC-VARIAN analyzer. For quality assurance,
one analytical blank was used for every 20 samples being considered
C. dos Santos Vergilio, D. Lacerda, T. da Silva Souza et al. Science of the Total Environment 756 (2021) 143697

Table 1
Physical and chemical parameters of the water and sediment of the sampling sites in the Doce River basin. SPM = Surface particulate matter. Significance codes: *** = 0.001; ** = 0.01;
* = 0.05; . = 0.1.

Sampling Damming distance (km) Sites Water Sediment

SPM pH Conductivity Dissolved Oxigen Turbidity Sand (%) Silt (%) Clay (%)
(mg·L−1) (μS·cm−1) (mg·L−1)

1 256 Ipatinga 1877 6.6 120 4.7 6750.0 55 45 0

365 Gov. Valadares 906 7.1 81 5.1 3843.0 14 71 15
407 Tumiritinga 1411 6.8 100 4.2 2630.0 8 66 26
476 Resplendor 806 6.8 100 4.4 6018.0 0 36 64
500 Aimorés 305 6.6 122 4.5 960.0 93 6 1
523 Mascarenhas 336 6.1 120 4.8 1079.0 100 0 0
556 Colatina 350 7.4 118 6.2 1179.0 100 0 0
622 Linhares 1070 7.6 107 4.3 2704.0 100 0 0
666 Regência 747 6.7 948 5.4 1447.0 86 14 0
2 2,5 Bento Rodrigues 166 7.5 117 5.4 13.5 54 32 14
71 Gualaxo 157 7.5 68 5.8 2.7 54 33 13
157 São José do Goiabal 95 7.5 82 5.4 44.0 48 37 15
242 Bom Jesus do Galho 120 7.5 69 5.7 52.9 44 40 17
256 Ipatinga 67 7.6 98 5.8 31.4 43 39 18
365 Gov. Valadares 14 7.1 87 6.2 11.3 46 36 18
407 Tumiritinga 19 7.6 91 5.0 14.0 42 39 20
476 Resplendor 14 7.4 98 5.0 12.1 38 40 22
500 Aimorés 9 7.9 73 5.2 14.0 43 37 20
523 Mascarenhas 13 7.9 41 7.0 17.8 60 26 14
556 Colatina 16 8.0 46 6.0 16.6 50 33 17
622 Linhares 19 8.1 89 6.1 17.4 33 44 23
666 Regência 16 7.8 2407 7.2 12.0 0 65 34
Mean (sampling 1) 868*** 6.8 202 4.8 2956*** 62 26 12
SD (sampling 1) 529 0.4 280 0.7 2169.0 43 29 21
Mean (sampling 2) 56 7.7• 259 5.8 20.0 43 38 19*
SD (sampling 2) 59 0.3 646 0.7 14.2 15 9 6

the results with the coefficient of variation was less than 15%. For SPM,
the analytical blanks contained glass fiber filters that passed through the
same procedures of acid digestion mentioned previously. To determine
the accuracy of the method, the standard reference material 1646a Estu-
arine Sediment from National Institute of Standards & Technology –
NIST was used following the same procedures mentioned above. The
quantification and the method's detection limits together with recovery
values of the certified material are demonstrated in Supplementary
Material S2.
in Erlenmeyer flasks to obtain exponential cultures. The growth inhibition
test was performed according to ABNT NBR 12648 (ABNT, 2018). For the
control (only with LC Oligo medium) and tested waters, three replicates
were prepared in sterile flasks. A cell concentration of 1 × 104 cells/mL
was added to each replicate. The solutions were maintained under contin-
uous agitation with white fluorescent light at 23 °C and 27 °C for 96 h. At
the end of the test, three aliquots each from the control and treated sam-
ples were examined for counting under an optical microscope with a
Neubauer chamber under a 100× objective.

2.3. Enrichment factor (EF) in sediment
The enrichment factor is frequently used in geochemical studies to
determine the degree of anthropogenic metal pollution:
 
Metal
sample
Fe
EF ¼  
Metal
background
Fe
Iron was used as the normalizing element because of its conservative
nature and low mobility, which reduces the effects of grain size and
mineralogy. Background values were considered as the median of
each element from the sediment samples from the Iron Quadrangle
region of Minas Gerais, the region where the dam is located (Vicq
et al., 2015). According to Sakan et al. (2009), the EF values are
interpreted as follows: EF < 1 indicates no enrichment; < 3 is minor
enrichment; 3–5 is moderate enrichment; 5–10 is moderately severe
enrichment; 10–25 is severe enrichment; 25–50 is very severe enrich-
ment; and > 50 is extremely severe enrichment.
2.4. Immediate effects (15 days) of Doce River to biological indicators
2.4.1. Algal toxicological assay (first trophic level)
Starter cultures of Raphidocelis subcapitata were prepared by inoculat-
ing a loop of algal cells (from an agar plate) in 100 mL of LC Oligo medium
2.4.2. Daphnia similis toxicological assay (second trophic level)
The acute toxicity tests were performed according to ABNT NBR
12713 (ABNT, 2016a), where eight replicates with five neonates 6 to
24 h old were exposed to 10 mL of water samples for 48 h under static
conditions at 20 ± 2 °C in the dark. The control samples were main-
tained only with MS medium. After exposure, immobilized organisms
were counted. Tests were considered acceptable if D. similis immobility
in the negative controls did not exceed 10%.
2.4.3. Danio rerio toxicological assay (third trophic level)
This study was performed in accordance with the ethical guidelines
and the experimental methods were approved by the Committee on
Ethical Used of Animals of Espírito Santo Federal University, Alegre
Campus (CEUA Alegre) with protocol number 26/2016. Danio rerio
specimens (zebrafish) were obtained from the Ornamental Fish Produc-
tion and Nutrition Laboratory (LNPEO) of the Federal Institute of
Espírito Santo (IFES) Campus Alegre. The fish were acclimated to the
dechlorinated water used in the tests, by replacing 25% of the water vol-
ume every 24 h. After the total replacement of the water, the organisms
were kept in this water for at least seven days before the beginning of
the tests. The acute toxicity tests were performed according to ABNT
NBR 15088 (ABNT, 2016b). Ten adult organisms (total length: 2.0 ±
1.0 cm) were exposed to the water samples for 96 h in a semi-static
test, with renewal of the test solution every 24 h. The observed effect
was the lethality in comparison to the control under the same condi-
tions. The test was repeated twice as experimental replicates.

4
C. dos Santos Vergilio, D. Lacerda, T. da Silva Souza et al.
2.4.4. Metal accumulation in fish
An aliquot of 0.2–0.5 g (wet weight) obtained from a pooled sample
(five fish of the same treatment) was solubilized in 65% HNO3 at 150 °C.
The metal determination (Al, Ba, Cd, Cr, Cu, Fe, Mn, Pb and Zn) was per-
formed in resuspended 0.5% v/v HNO3 extracts with inductively coupled
plasma-optical emission spectrometry (ICP-OES Varian - 720 ES). To de-
termine the accuracy of the method, the standard reference material
DORM-3 fish protein reference material from National Research Council
Canada was used following the same procedures mentioned above. The
elements evaluated (exception of Ba) in fish muscle were choose based
on recovery (80–120%) values of DORM 3. Ba was also included in the
analysis, since the values were comparable to other published studies
with metal concentration in fish muscle tissue using the same methodol-
ogy (Agostinho et al., 2020; Lacerda et al., 2020). The quantification and
the method's detection limits are presented in Supplementary Table S2.
2.5. Immediate (15 days) and long term (six months) toxicogenetic poten-
tial of Doce River water and sediment
Allium cepa seeds (bundle-branch variety) from the same lot (num-
ber 774758, 90% germination, Isla®) were used for being genetically
and physiologically homogeneous (Leme and Marin-Morales, 2009).
The seeds were germinated in Petri dishes with filter paper embedded
in the water or in the whole sediment collected from the different sam-
pling points. Distilled water was used as negative control and 0.025%
colchicine as positive control. After five days, A. cepa (1.5–2 cm) rootlets
were collected and fixed in methanol + acetic acid in the ratio 3: 1.
Next, the rootlets were hydrolyzed in 1 N HCl at 60 °C for 10 min and
then subjected to Schiff's reagent for 2 h in the dark. The root tips (mer-
istematic region) were sectioned, stained with 2% acetic carmine, cov-
ered by coverslip and macerated. The slides were immersed in liquid
nitrogen and then mounted with Canada balsam. The material was an-
alyzed under a light microscope with magnification of 400×. For each
sampling point, 5000 cells were counted (500 cells in 10 slides). Cyto-
toxicity of the Doce River samples was determined by the mitotic
index (cell division/total cells observed x 100). Aberrant cells, observed
in each state of the cell cycle, were grouped into the same category by
the genotoxicity index. Micronuclei were counted to determine the mu-
tagenic potential of water and sediment samples.
2.6. Statistical analysis
The data on the differences of metal concentrations (dissolved, SPM,
sediment and fish) and the physical-chemical values evaluated in the
sampling campaigns were submitted to ANOVA (aov, base package, R
Core Team, 2020), assuming a 95% confidence level, using the R soft-
ware (R Core Team, 2020). Data were transformed to meet the assump-
tions of ANOVA (normality, linearity and homoscedasticity of residuals)
using a maximum likelihood function (boxcox, MASS package, Venables
and Venables and Ripley, 2002). Since the normality criteria were not
met by the R. subcapitata, D. similis and A. cepa data, the Kruskal-
Wallis nonparametric test (Kruskal, agricolae, Mendiburu, 2019) was
applied assuming a 95% confidence level.
To evaluate metal associations and behavior in SPM and sediment
along the rivers, Pearson's correlation coefficients were calculated to iden-
tify clusters of metals with the same overall profiles regardless of their
concentrations at each sampling site, using the R software (cor, base pack-
age, R Core Team, 2020). To conduct this analysis, the variables with miss-
ing data (less than 50% of missing data) were regressed with other
variables and the values were stipulated using linear regression with R2
greater than 0.80. The clusters were defined with pairwise variables
that presented a strong correlation coefficient (greater than 0.70). The
tendencies of metal concentrations were plotted according the associa-
tions verified in the clusters, using the R software (R Core Team, 2020)
with data transformed to Z-scale (standard scale in statistics where the
standard deviation is 1 unit. and the mean is zero), to make the variables
5
Science of the Total Environment 756 (2021) 143697
comparable. As the quantified elements in the dissolved fraction showed
a low correlation with each other, the graphical analysis of the element's
behavior was performed only for SPM and sediment.
The cytogenotoxicity results in water and sediments were associated
with metal groups through of Pearson's correlation (cor.test, base pack-
age, R Core Team, 2020). The association between genotoxicity and the
mutagenicity with mitotic index was evaluated by ANCOVA (lm, base
package, R Core Team, 2020), assuming a 95% confidence level. The
data were transformed to meet the assumptions of ANOVA and
ANCOVA (normality, linearity and homoscedasticity of residuals)
using a maximum likelihood function (boxcox, MASS package, Venables
and Venables and Ripley, 2002).
3. Results
3.1. Elements in the dissolved and suspended particulate matter
With the dam rupture (sampling 1), the tailings released were
mainly transported as SPM, raising the turbidity levels (Table 1). After
the dam rupture (sampling 1–15 days) the SPM showed the predomi-
nance of Fe (mean value: 121.7 mg·L−1), Cr (mean value: 91.05 mg·L−1),
Al (mean value: 64.60 mg·L−1), Cu (mean value: 24.92 mg·L−1), Pb
(mean value: 20.84 mg·L−1), Cd (mean value: 14.10 mg·L−1), together
with the rare earth metals La (mean value: 184.99 mg·L−1), Y (mean
value: 74.24 mg·L−1), Gd (mean value: 71.92 mg·L−1), and Nd (mean
value: 66.35 mg·L−1). Six months later (sampling 2), the elements
showed significant decrease in their levels, but Fe (mean value:
18.10 mg·L−1), Cr (mean value: 10.52 mg·L−1) and Al (mean value:
9.2 mg·L−1) continued to be the majority elements in SPM, together
with Nd (mean value: 19.21 mg·L−1).
The SPM acted in the metal transport along the river, since increased
levels of most of the metals were observed in this matrix (Fig. 2a - Group
1 / 15 days: Fe, Al, Ti, Mn, Zn, Cr, Ni, Cu, Pb, Cd, Eu, Gd, La, Lu, Pr, Sc, Tb, Y,
Supplementary Material S3), especially closest to the rupture site
(Ipatinga – 256 m) (Fig. 2a – Group 1 / 15 days). However, the existence
of four water dams (for electricity generation), which were closed after
the dam rupture, acted to retain the material transport along the river,
reducing the SPM and turbidity (Table 1) together with the metal levels
at sampling sites downstream from the water dams (Governador
Valadares – 365 km, Aimorés – 500 km and Mascarenhas – 523 km)
(Fig. 2a – Group 1 / 15 days). An increase of the metal concentrations
of this group near of the river mouth (Regência – 666 m) (Fig. 2a) oc-
curred together with increases of conductivity, SPM and silt levels
(Table 1), which indicates the existence of a mixture zone due to the
meeting of the river with the Atlantic Ocean.
Six months later (sampling 2), less SPM was being transported
along the river, causing lower turbidity (Table 1), reducing the
levels of these metals (Fig. 2a - six months, Supplementary Mate-
rial S3). However, there was still a trend of higher metal concentra-
tions near the dam (Bento Rodrigues - 2.5 km, Gualaxo – 71 km, São
José do Goiabal – 157 km, Bom Jesus do Galho – 242 km and
Ipatinga – 256 km), together with increased levels of SPM and tur-
bidity (Table 1). This occurred because, despite attempts, the total
blockage of the tailings was not successful, so part of the tailings
was still slowly carried by the river. Besides that, the increase of
rainfall in the upstream part of the watershed might also have
acted to carry away material deposited on the banks of the river
(Fig. 2a).
In relation to the dissolved elements, some metals in the dis-
solved fraction showed concentrations below the method's detec-
tion limit in both sampling campaigns (Cd, Cu, Er, Gd, Lu, Ni, Pb,
Tm, Yb). From the quantified elements, the majority (Al, Ba, Ce, Cr,
Eu, Fe, La, Mn, Nd, Pr, Sc, Sm, Tb, Ti, Y and Zn) presented higher con-
centrations in the immediate moment (15 days) after the dam rup-
ture with a decrease in their levels 6 months latter (Supplementary
Material S4).
C. dos Santos Vergilio, D. Lacerda, T. da Silva Souza et al. Science of the Total Environment 756 (2021) 143697

Fig. 2. Behavior of metal levels in (a) suspended particulate matter (SPM) and (b – d) sediment in the period immediately after (15 days) (brown lines) and six months after (blue lines) of
the tailings dam rupture along the Doce River. (a) Group 1: Fe, Al, Ti, Mn, Cr, Ni, Cu, Pb, Cd, Eu, Gd, La, Lu, Pr, Sc, Tb and Y, Group 2: Se, Ba, Zn, Er, Nd and Sm, All elements: Fe, Al, Ti, Mn, Se, Ba,
Zn, Cr, Ni, Cu, Pb, Cd, Ce, Eu, Gd, La, Lu, Nd, Pr, Sc, Sm and Tb. (b) All elements: Fe, Al, Ti, Mn, Se, Ba, Zn, Cr, Ni, Cu, Pb, Cd, Hg, Er, Eu, Gd, La, Lu, Nd, Pr, Sc, Sm, Tb and Y, Major group: Fe, Al, Ti,
Mn, Ba, Zn, Cr, Ni, Cu, Pb, Cd, Hg, Ce, Er, Gd, La, Lu, Nd, Pr, Sc, Sm and Tb. The values were transformed to z-scale to make them comparable. The dashed lines represent four water dams along
the river. The bars represent the standard error of z-scaled variables at each sampling site.

3.2. Elements in the sediments
After the dam rupture (sampling 1–15 days) the sediment showed
the predominance of Fe (mean value: 39.02 mg·g−1), Al (mean
value: 23.68 mg·g−1), Ti (mean value: 1.07 mg·g−1), Mn (mean
value: 0.46 mg·g−1), Se (mean value: 85 μg·g−1), Ba (mean value:
56 μg·g−1), Zn (mean value: 29 μg·g−1), Cr (mean value: 30 μg·g−1),
Ni (mean value: 13 μg·g−1), Cu (mean value: 12 μg·g−1) and Pb
(mean value: 7.16 μg·g−1). Six months latter (sampling 2), most of
elements showed increase in their levels, but the Al (mean value:
167.98 mg·g−1), Fe (mean value: 109.66 mg·g−1), Ti (mean value:
1.97 mg·g−1), Mn (mean value: 0.79 mg·g−1), continued to be the
majority elements in the sediment, together with elements Ba
(mean value: 167.98 μg·g−1), Hg (mean value: 0.10 μg·g−1), Pb
(mean value: 10.78 μg·g−1), Tb (mean value: 58.93 μg·g−1) and Er
(mean value: 2.87 μg·g−1) hat showed significant increase in their
concentrations six months after the dam rupture.
In the sediments there was an increase of the metal levels six
months (sampling 2) after the dam rupture in relation to the first sam-
pling (15 days) (Fig. 2b, Supplementary Material S5). In both samplings,
the highest metal levels in sediments were observed nearest the mining
source in Bento Rodrigues – 2.5 km (Fig. 2b). The multi-elemental anal-
ysis demonstrated the same behavior for most of the elements 15 days
after the dam rupture (sampling 1) (Fig. 2b – 15 days), while six months
later, the Ba and Mn (Fig. 2c) and Ce, Y, Er, Nd, Sm, La (Fig. 2d) formed
groups with distinct behaviors. At Resplendor – 476 km and Aimorés
– 500 km, the increase of metal concentrations in the sediment (sam-
plings 1 and 2) was influenced by their upstream position to a water
dam. The consequent blockage of the SPM induced the metal transport
to the sediment. On the other hand, in the sampling points downstream
from the water dams (São José do Goiabal – 157 km, Governador
Valadares – 365 km, Tumiritinga – 407 km, Mascarenhas – 523 km
and Colatina – 556 km), a decrease in metal concentration was observed
(Fig. 2b). The levels of Cr, Ni, Cd and Hg in sediment were higher than
the threshold effects level (TEL) established by the National Oceanic
and Atmospheric Administration (NOAA). Concentrations above the
PEL (probable effects levels) were observed for Ni and Cd. The highest
incidence of non-conforming values was observed six months after
the dam rupture (sampling 2) (Supplementary Material S5).
3.3. Enrichment of metal levels along the Doce River
The elements Cd, Ti, Al, Zn, Pb and Ni had the highest enrichment
factors (moderately severe enrichment or even very severe enrich-
ment) in the period immediately after (15 days) the Fundão Dam
rupture (Table 2). The metal enrichment was more evident promptly

6
C. dos Santos Vergilio, D. Lacerda, T. da Silva Souza et al. Science of the Total Environment 756 (2021) 143697

Table 2
Enrichment factors (EFs) from the sediment samples along the Doce River. The values and colors indicate: <1 no enrichment, <3 minor enrichment, 3–5 moderate
enrichment, 5–10 moderately severe enrichment, 10–25 severe enrichment, 25–50 very severe enrichment.

Damming
Sampling Sites distance Fe Al Ti Mn Zn Cr Ni Cu Pb Cd
(km)
1 Ipatinga 256 1 7.3 24.8 2.9 2.7 1.9 3.0 2.2 1.1 13.9 <1
Gov. Valadares 365 1 5.7 39.4 1.9 7.1 2.0 3.3 3.1 2.5 9.8 <3
Tumiritinga 407 1 4.3 31.3 6.5 4.8 2.1 4.0 2.8 1.4 10.1 3-8
Resplendor 476 1 3.9 34.7 2.5 4.1 3.7 3.8 2.5 1.0 8.1 5 - 10
Aimorés 500 1 3.1 3.8 1.1 1.2 1.0 1.6 1.6 0.8 23.5 10 - 25
Mascarenhas 523 1 4.6 6.5 2.0 1.9 1.3 2.3 2.3 1.1 22.1 25 - 50
Colatina 556 1 4.9 46.3 1.3 2.5 2.0 2.7 1.9 3.9 17.0
Linhares 622 1 2.8 9.2 0.8 1.3 0.7 1.7 1.0 0.3 18.1
Regência 666 1 3.7 5.4 2.0 2.0 1.0 1.8 2.6 0.8 25.4
2 Bento Rodrigues 3 1 5.2 4.6 0.7 2.1 1.0 1.6 1.9 1.7 10.3
Gualaxo 71 1 9.3 30.1 1.9 3.0 2.0 2.7 1.9 3.6 13.9
São José do Goiabal 157 1 16.8 20.1 2.4 5.7 2.9 4.0 3.7 5.9 26.0
Bom Jesus do Galho 242 1 21.5 25.4 2.2 5.3 4.6 6.0 4.9 8.3 38.6
Ipatinga 256 1 1.0 4.1 0.5 0.7 0.6 0.8 0.6 0.5 2.5
Gov. Valadares 365 1 1.3 5.3 0.7 0.8 0.7 0.8 0.7 0.6 2.8
Tumiritinga 407 1 1.8 11.6 1.1 1.7 1.2 1.7 1.4 0.8 3.5
Resplendor 476 1 4.8 26.0 1.5 2.1 1.4 1.6 1.4 1.8 7.6
Aimorés 500 1 0.8 3.2 0.7 1.1 0.7 0.8 0.5 0.4 3.6
Mascarenhas 523 1 1.4 5.6 0.6 0.9 0.7 1.1 0.6 0.2 3.2
Colatina 556 1 6.4 10.3 5.3 2.5 3.1 3.4 3.0 2.3 14.5
Linhares 622 1 1.3 1.5 4.1 0.4 0.4 0.8 0.5 0.2 3.5
Regência 666 1 0.5 0.4 0.3 0.3 0.2 0.2 0.1 0.2 3.9

after the dam rupture, with a tendency to decrease until six months
later (Table 2).
3.4. Immediate biological effects of the tailings
The tailings had immediate impacts (after 15 days) on different
levels of the trophic chain of producers - algae, primary consumers -
microcrustaceans (Fig. 3a, b). Growth was inhibited of the
chlorophyceae alga Raphidocelis subcapitata – indicative of the first tro-
phic level (Fig. 3a) – after the exposure to water from almost all sam-
pling sites of the Doce River (with exception of Governador Valadares
– 365 km and Tumiritinga – 405 km). Immobility of the microcrustacean
Daphnia similis – indicative of the second trophic level (Fig. 3b) – was
observed from the closest site to the dam (Ipatinga – 256 km) until
Mascarenhas – 523 km (with exception of Aimores – 500 km). The
Doce River water in the period immediately after (15 days) the dam
rupture did not cause significant mortality of the Danio rerio specimens
(Fig. 3c).
3.5. Immediate and long-term metal accumulation by fish
Although without mortality, the fish exposed to the water of the
Doce River accumulated Al, Fe, Mn and Zn in the muscle tissue
(Fig. 4). Particularly for Mn and Zn, the accumulation increased with
the exposure to the water six months after the dam rupture (Fig. 4).
In contrast, for Ba, Cr and Cu, the accumulation effect was not observed
(Fig. 4). Cd and Pb levels were below the method's detection limits for
both sampling periods.
3.6. Immediate and long-term toxicogenetic effects
Both water (with high amount of SPM) and sediment from the Doce
River had cytotoxic, genotoxic and mutagenic effects in the period im-
mediately after (15 days) and six months after the dam rupture
(Fig. 5). After 15 days (sampling 1), the water from all sampling sites
demonstrated cytotoxicity (Fig. 5a) and mutagenicity (Fig. 5c) (with ex-
ception of Ipatinga – 256 km). The water genotoxicity was also evident
along the Doce River (Fig. 5b) with the exception of Ipatinga – 256 km
and in sampling sites close to the mouth (Colatina – 556 km and
Regência – 666 km). The absence of genotoxicity and mutagenicity at
Ipatinga – 256 km occurred due to the higher cytotoxic effect. Moreover,
the highest cytotoxicity, genotoxicity and mutagenicity were observed
at Aimorés (500 km) (Fig. 5a - c), due to the blockage effect of the
dam, which acted to increase the levels of the elements in the water
(associated with the SPM). Six months later (sampling 2), water cyto-
toxicity was still observed near the dam (Bento Rodrigues – 2.5 km
and Ipatinga – 256 km) (Fig. 5a). Compared to the sampling 1
(15 days), the frequency of genetic damage and number of micronuclei
increased considerably, mainly due to the genotoxic (Fig. 5b) and muta-
genic effects (with exception of Bento Rodrigues – 2.5 km due to higher
cytotoxic effect) (Fig. 5b) along the entire river course.
In relation to the sediment, the samples collected shortly after the
rupture (15 days) along all the river course were extremely cytotoxic
(Fig. 5d), since the exposure induced arrest of meristematic cells in
the interphase, due to the interruption of the cell division cycle. Conse-
quently, these samples were neither genotoxic nor mutagenic. Six
months later, although cytotoxicity was no longer observed in the
sediment (Fig. 5d), the genotoxicity was evident in all sampling sites.
Micronucleus frequency was significantly higher only at Governador
Valadares - 365 km from the dam rupture site (Fig. 5f).
The covariance analysis between cytogenotoxic variables demon-
strated a negative or absent association between genotoxic abnormali-
ties and mitotic index in the period immediately after the dam
rupture (sampling 1–15 days) in water and sediment respectively. On
the other hand, a positive association was noted between genotoxicity
and mitotic index in both water and sediment after six months
(Fig. 6a). The mutagenicity (indicated by the prevalence of micronuclei)
was positively associated with the mitotic index in both sampling
periods and matrices (Fig. 6b).
The associations mentioned above influenced the correlations
between cytogenotoxic data with the metal groups in SPM and sedi-
ment in both sampling periods. The mitotic index and micronucleus
rate showed mostly a negative correlation with the elements present
in the SPM 15 days and six months after the rupture. This indicates
that the higher level of the elements in SPM caused a decrease in the
mitotic index, which consequently influenced the micronucleus

7
C. dos Santos Vergilio, D. Lacerda, T. da Silva Souza et al.
Fig. 3. Toxicology tests with representative species of different trophic levels exposed to
the water of the Doce River after 15 days after the dam rupture. (a) Chlorophyceae
algae – Raphidocelis subcapitata (primary producer – first trophic level),
(b) microcrustacean – Daphnia similis (primary consumer – second trophic level), and
(c) fish – Danio rerio (secondary consumer – third trophic level). NC: Negative control.
The dashed lines represent the four water dams along the river. The letters represent
statistically different values according to the Kruskal-Wallis test (p < 0.05) and compare
the different effects among the sampling sites. * means significant compared to the
negative control. The bars represent the standard errors.
incidence (Table 3). In the sediments, there was a positive correlation of
the mitotic index with all elements quantified 15 days after the dam
rupture. On the other hand, there was a negative and significant
8
Science of the Total Environment 756 (2021) 143697
Fig. 4. Metal (Al, Ba, Cr, Cu, Fe, Mn and Zn) accumulation in the muscle of Danio rerio fish
exposed to Doce River water from the immediate moment (15 days) and six months after
the tailings dam rupture. NC: Negative control. The letters represent statistically different
values according the Tukey test (p < 0.05) and compare the treatments (NC, 15 days and
six months) for each element. The distance between y-axis values in the graph was log-
transformed to optimize data visualization.
association of most of the elements (Fe, Al, Gd, Lu, Ti, Zn, Cr, Ni, Cu, Pb,
Cd, Hg, Pr, Sc, Tb and Mn) with the mitotic index, genotoxic abnormal-
ities and micronucleus rate. Those elements included all that were
above the TEL and PEL (Cr, Ni, Cd and Hg), while other elements were
positively correlated with those variables (i.e., Ce, Y, La and Ba)
(Table 3). These data clearly indicate of the presence of one group of el-
ements that was more correlated with severe adverse toxicogenetic ef-
fects (Table 3).
4. Discussion
After the dam rupture, the fine particulate matter of the tailings in-
duced an increase in turbidity levels and metal distribution along the
whole course of the Doce River until the Atlantic Ocean. Therefore, the
SPM was a relevant environmental matrix for the dispersion of metals
along the river until its mouth (Hatje et al., 2017). The present study
demonstrates that in the long term (six months after the rupture), the
SPM settled as bottom sediment, increasing the metal levels together
with the potential to induce biological and toxicogenetic damages.
The SPM was at least two orders of magnitude higher than the his-
toric average (51 mg/L) in the month of the dam rupture (IGAM,
2015). This was also observed even three months afterward (Hatje
et al., 2017). SPM levels are affected by the particles and organic matter
loads from the surrounding soils. This surface runoff is increased during
the rainy season leading to an increase in SPM levels. The time series of
monthly rainfall of the Doce River basin shows a clear division between
dry and rainy periods (Coimbra et al., 2019), being the rainy season re-
sponsible for approximately 94% of the SPM supply (Oliveira and da
Silva Quaresma, 2017). Therefore, it was already expected seasonal dif-
ferences in SPM levels together with metal concentrations between the
samplings. However, in 2015, the Doce River experienced an extreme
drought. The median of the Doce River discharge in November 2015
(~320 m3/s) was well below historically median values for November
(790 m3/s) (Oliveira and da Silva Quaresma, 2017). Consequently, the
fluctuations in SPM levels together with metals concentrations ob-
served in the present study occurred mainly due to dam rupture. This
C. dos Santos Vergilio, D. Lacerda, T. da Silva Souza et al. Science of the Total Environment 756 (2021) 143697

a ab a
ab ab ab
bc
NC bc
NC
PC bc*
cd* bcd PC
cd*
ef* de*
f* f*
f*
ef*

g* g* g* g*

a*
a*
PC PC
a*

b*
bc* bc* b* bc*
bc*
d* cd*
e*
fg ef* fg hij
NC ghij NC fgh fghi j j ij

a*

ab*
PC PC
cde
b*
bcd*
bcde* bcde* NC
cde*
cde* bc* cde de
de cde de
e e e
NC e e e e

Fig. 5. Cytotoxicity (a and d), genotoxicity (b and e) and mutagenicity (c and f) of Doce River water (a, b and c) (blue lines) and sediment (d, e and f) (brown lines) in the period
immediately after (15 days) and six months after the tailings dam rupture. NC: Negative control. PC: Positive control. The dashed lines represent the four water dams along the river.
The letters represent statistically different values according the Kruskal-Wallis test (p < 0.05) and compare the different treatments, sampling period and exposed matrices altogether
for each variable. * means significant compared to the negative control.

observation is in accordance with other studies that demonstrated that
the suspended load of the river was extremely high due to the dam tail-
ings (Coimbra et al., 2019; Quaresma et al., 2020). Therefore, the de-
crease in SPM and metal concentrations observed six months later of
the dam rupture, demonstrated the precipitation of the SPM that con-
solidated the bottom sediment, together with transport of the material
to the river mouth.
The material transport along the river course is also demonstrated
through differences in the sediment particle size composition. In first
sampling, the sampling sites close to the river mount (Colatina –
556 km, Linhares – 622 km and Regência – 666 km) were totally
sandy. While, six months later the increase in silt and clay particles to
the sediment composition was evident along the river course, changing
the granulometry until the sites close to the sea (Table 1), evidencing
the SPM deposition along the Doce River.
SPM transport and deposition along the Doce river might also result
in the reduction of the dissolved elements in the water column ob-
served 6 months after the dam rupture. Since the majority of the dis-
solved elements (Al, Ba, Ce, Cr, Eu, Fe, La, Mn, Nd, Pr, Sc, Sm, Tb, Ti, Y
and Zn) presented higher concentrations only in the immediate mo-
ment (15 days) after the dam. Our results are in accordance with
Viana et al. (2020) that showed increase in the concentrations of dis-
solved Fe, Al, Ba, and Zn after the dam rupture in comparison with pre-
vious levels. According to Hatje et al. (2017) the dissolved Fe, Ba, Cu, Ni,

9
C. dos Santos Vergilio, D. Lacerda, T. da Silva Souza et al. Science of the Total Environment 756 (2021) 143697

Fig. 6. Association between the (a) genotoxic abnormalities and the mitotic index and (b) the micronuclei and the mitotic index from water and sediment samples from the immediate
moment (15 days) after the dam rupture. The shadings identify the 68% confidence interval of the regression models.

Cr, Th and Al presented increase in transport and flux peaks along the
Doce river basin. Therefore, the material transport along the river asso-
ciated with processes, such as flocculation, adsorption onto SPM, and
coagulation in the water column might be involved in the removal of
these elements from water column into the bottom sediment.
However, until total precipitation and due to remobilization events,
these elements can be released into water column, where they accumu-
late through biota and become incorporated through the food chain.
Therefore, studies of the metal concentrations with the local biota or
with bioassays are important to demonstrate toxicological conse-
quences to different animal species, together with the evaluation of
the risks to humans. The present study demonstrates that the water
(15 days after the dam rupture) with high amount of SPM was able to
induce immediate impact at different trophic levels, from algae to
microcrustaceans and fish. Indeed, the fish's exposure to the Doce
River water from the immediate period (15 days) and long-term (six
months) after the dam rupture led to the accumulation of Al, Fe, Mn
and Zn in the muscle tissue. This is an alert about the effects in the ex-
posed biota beyond the risk of consuming fishery items by the local
population.
After the dam rupture (sampling 1–15 days) the sediment showed
the predominance of Fe (mean value: 39.02 mg·g−1), Al (mean
value: 23.68 mg·g−1), Ti (mean value: 1.07 mg·g−1), Mn (mean
value: 0.46 mg·g−1), Se (mean value: 85 μg·g−1), Ba (mean value:
56 μg·g−1), Zn (mean value: 29 μg·g−1), Cr (mean value: 30 μg·g−1),
Ni (mean value: 13 μg·g−1), Cu (mean value: 12 μg·g−1) and Pb
(mean value: 7.16 μg·g−1). The predominance of Fe, Mn, Cr, Zn, Ni, Cu
and Pb was also found to be correlated with the tailings characteristics,
composed of Fe-oxyhydroxides (goethite, hematite), kaolinite and
quartz (Queiroz et al., 2018). The dominance of these minerals is associ-
ated with the characteristics of the mined ore (e.g., itabirite rocks) (Silva
et al., 2018).
Cd, Ti, Al, Zn, Pb and Ni enrichment together with Cr, Ni, Cd and Hg
levels beyond TEL and PEL also increase the importance of monitoring
programs, including not only the river abiotic matrices but also the
biota. The accumulation of these elements in organisms can cause neg-
ative effects on the ecosystem and human health through ingestion of
contaminated food.
Besides metals characteristic of the tailings content, some rare ele-
ments or those with normally low levels also had high levels after the
dam rupture in SPM (Gd, La, Lu, Nd, Sm, Tb, Y) and in sediment (La
and Y). The northern region of Rio de Janeiro and Espírito Santo states
presents anomalies of the mineral monazite, phosphate-containing
metals and rare earths - an important source of Th, La and Ce (Carmo
et al., 2018). Therefore, in light of the local soil composition, the mining
activity might be acting to increase these metals in the tailings that were
released with the dam rupture. Rare earth elements are considered a
concern, especially from mining and industrial activities, due to their
persistent toxicity, radioactive nature, and potential for bioaccumula-
tion (Khan et al., 2016). Beyond that, many of their toxicological effects
are unknown. Indeed, the Fundão Dam rupture increased the concen-
tration of the rare earth metals in the SPM and sediment. These

Table 3
Pearson's correlation of cytogenotoxicity variables obtained in the Allium cepa test with bioavailable metal groups. * statistically significant (p < 0.10); ** statistically significant (p < 0.05).

Matrice Sampling Element groups Mitotic index Genotoxic abnormalities Micronuclei

SPM 1 Fe. Al. Ti. Mn. Cr. Ni. Cu. Pb. Cd. Eu. Gd. La. Lu. Pr. Sc. Tb and Y −0.51 0.26 −0.54
Se. Ba. Zn. Er. Nd and Sm −0.65 0.30 −0.40
2 Fe. Al. Ti. Mn. Se. Ba. Zn. Cr. Ni. Cu. Pb. Cd. Ce. Eu. Gd. La. Lu. Nd. Pr. Sc. Sm and Tb −0.93** −0.66 −0.49
Sediment 1 Fe. Al. Ti. Mn. Se. Ba. Zn. Cr. Ni. Cu. Pb. Cd. Hg. Er. Eu. Gd. La. Lu. Nd. Pr. Sc. Sm. Tb and Y 0.55 0.20 –
2 Fe. Al. Gd. Lu. Ti. Zn. Cr. Ni. Cu. Pb. Cd. Hg. Pr. Sc and Tb −0.86** −0.56 −0.30
Nd and Sm 0.73* 0.68 0.68
Ce and Y 0.55 0.30 0.80*
La 0.54 0.43 0.02
Mn −0.76* −0.40 −0.26
Ba 0.51 0.59 0.81**
Er −0.43 0.02 0.43

10
C. dos Santos Vergilio, D. Lacerda, T. da Silva Souza et al.
elements are now in the aquatic system so their levels should be mon-
itored, together with the possibility of accumulation through local biota.
Our results also demonstrated that the water (with large amount of
SPM) and sediment along the entire course of the Doce River even six
months after the dam rupture had cytotoxic, genotoxic and mutagenic
potential. Preliminary analysis of the chemical and toxicogenetic analy-
sis of the tailings collected in Bento Rodrigues six days after the dam's
collapse already demonstrated the higher mobility potential of Al, Ba,
Fe, Mn, Pb, and Sr, as well as the potential for cytogenotoxicity and
DNA damage (Segura et al., 2016). Our results are also in accordance
with Quadra et al. (2019), who documented that water samples from
the Ipatinga River, collected a few days after the accident, altered
A. cepa cell cycle (decreased mitotic index and increased frequency of
aberrant cells). However, in the present study, we found that even six
months after the dam rupture, the water and sediment samples from
the entire course of the Doce River showed toxicogenic potential. This
means that despite the decrease in SPM, turbidity, and metal levels
along the Doce River due to deposition or the blockage effect of the
water dams, these effects are not enough to mitigate the cytogenotoxic
action of the tailings, which was detected until the river mouth, more
than 666 km from the rupture site. These data indicate the need for
long-term monitoring along the entire Doce River course.
Sediment was a sensitive matrix in the evaluation of the toxicogenic
damage after the dam rupture. After that event, the river carried a con-
siderable volume of tailings, much of which was deposited in the bot-
tom sediment. After 15 days, severe cytotoxicity was already evident
in the sediment samples along the entire course of the Doce River.
This mitodepressive action resulted in the low rate of aberrant cells
and micronuclei. Therefore, mitotic index analysis is an important tool
to characterize the toxicity of contaminants (and environmental matri-
ces) because when an agent is highly cytotoxic, its genotoxic effect can
be masked (Hoshina and Marin-Morales, 2009; Dhyèvre et al., 2014,
Souza, Souza et al., 2018).
Finally, the present study demonstrates that the water dams along
the Doce River had important action to block SPM, inducing precipita-
tion to the sediment. Even after six months, the SPM and sediment levels
showed differences between the sampling points close to the dam until
upstream Aimorés – 500 km and Mascarenhas – 523 km in comparison
with the sampling points close to the river mouth. These data can direct
future environmental monitoring studies, considering the metal fluctu-
ations in the analyzed abiotic matrices along the Doce River.
5. Conclusions
The present study demonstrated the immediate and long-term
(after six months) impact of the Fundão tailings dam rupture on metal
concentration together with the biological and toxicogenic potential of
the water (with large amount of SPM) and sediment. Suspended partic-
ulate matter was an important geochemical support for metal transport
just after the rupture, and with time this material was transported to the
mouth and into the ocean, or underwent precipitation by oxy and hydro
colloidal iron complexes, increasing the metal levels in sediment. The
present results also show the negative effects of the tailings on different
levels of the trophic chain, together with the cytotoxic, genotoxic and
mutagenic potential of the water (with large amount of SPM) and sed-
iment along the entire course of the Doce River even six months after
the dam rupture. Long-term monitoring involving the abiotic and biotic
matrices is important to clarify the real impacts on the affected ecosys-
tems, and the contamination risks to human populations, especially be-
cause some toxic metals (Cr, Ni, Cd and Hg) presented levels above the
TEL and PEL (for Ni and Cd), implying the possibility of causing negative
effects on biota. Finally, much can be learned from this event. The
knowledge of the amplitude and long-term consequences is important
for future local actions. The learning acquired from the past events can
help to adopt proactive strategies to minimize social and environmental
impacts of future dam failure episodes.
11
Science of the Total Environment 756 (2021) 143697
Supplementary data to this article can be found online at https://doi.
org/10.1016/j.scitotenv.2020.143697.
CRediT authorship contribution statement
Cristiane dos Santos Vergilio: Conceptualization, Investigation,
Resources, Writing - original draft, Writing - review & editing, Visualiza-
tion, Supervision, Project administration. Diego Lacerda: Writing
- review & editing, Formal analysis, Data curation, Visualization. Tatiana
da Silva Souza: Investigation, Methodology, Supervision. Braulio
Cherene Vaz de Oliveira: Methodology, Validation, Data curation.
Vinicius Sartori Fioresi: Methodology. Victor Ventura de Souza: Meth-
odology. Giovana da Rocha Rodrigues: Methodology. Marjore Kinaip
de Araujo Moreira Barbosa: Methodology. Echily Sartori: Methodol-
ogy. Thiago Pessanha Rangel: Methodology, Validation. Diogo Quitete
Ribeiro de Almeida: Methodology, Validation. Marcelo Gomes de
Almeida: Methodology, Validation. Fabiano Thompson: Writing -
review & editing. Carlos Eduardo de Rezende: Conceptualization,
Investigation, Resources, Writing - review & editing, Visualization,
Supervision, Project administration.
Declaration of competing interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influ-
ence the work reported in this paper.
Acknowledgments
The authors are grateful for the support from the Laboratório de
Ciências Ambientais (LCA) and to the Programa de Pós-Graduação em
Ecologia e Recursos Naturais from the Universidade Estadual do Norte
Fluminense Darcy Ribeiro, the Laboratório de Morfologia Animal, the
Laboratório de Ecotoxicologia (LABTOX) and the Laboratório de Estudos
em Ciências Ambientais (LECA) from the Universidade Federal do
Espírito Santo – Campus Alegre for use of the infrastructure. The authors
thank the financial support from FAPES, CNPq (305217/2017-8), FAPERJ
(E-26/202.916/2017 and E-26/210.064/2018) and CAPES - Finance Code
001. Diego Lacerda received scholarship from FAPERJ (E-26/200.131/
2017).
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