REVISTA DE BIOLOGIA E CIÊNCIAS DA TERRA ISSN 1519-5228 Volume 24 - Número 2 - 2º Semestre 2024

1. REVISTA DE BIOLOGIA E CIÊNCIAS DA TERRA ISSN 1519-5228
Volume 24 - Número 2 - 2º Semestre 2024
BIORREMEDIAÇÃO POR BIOFILME COMO FERRAMENTA DE DESPOLUIÇÃO
AMBIENTAL EM ECOSSISTEMA LÓTICO
Flávia Martins Franco de Oliveira¹ & Maria Cristina Crispim2
RESUMO
Os ecossistemas aquáticos estão cada vez mais poluídos, principalmente os urbanos. É possível
aumentar a capacidade de autodepuração como ferramenta de despoluição aquática? O objetivo do
trabalho foi testar o aumento do biofilme como forma de aumentar a capacidade de autodepuração
em um rio tropical urbano poluído utilizando a biorremediação através da utilização do biofilme e
manejo de macrófitas (BioMac) na Paraíba – Brasil. Foram realizadas análises físicas, químicas e
biológicas e construídas estruturas de fixação para o biofilme, sendo um processo de biorremediação
por bioestimulação com a oferta de habitat. Os resultados mostraram, que após a inserção do
biotratamento houve uma grande diminuição dos nutrientes ao longo do rio, permitindo um aumento
da transparência, da diversidade biológica e o aumento do oxigênio dissolvido. Esse efeito foi mais
forte nos locais mais poluídos, como o Alto Rio do Cabelo que recebe esgotos. Conclui-se que o
sistema piloto de biotratamento com biofilme, foi eficaz e de baixo custo, tendo diminuído os
compostos fosfatados, aumentado o processo de nitrificação, o oxigênio, a transparência da água e a
diversidade de plantas, podendo ser utilizado em rios que recebem elevada carga de esgoto, como em
ambientes urbanos.
Palavras-chave: Biotecnologia, Biotratamento, Perifíton, Macrófitas.
BIORREMEDIATION BY BIOFILM AS AN ENVIROMENTAL DEPOLLUTION TOOL
IN LOTIC ECOSYSTEM
ABSTRACT
Aquatic ecosystems are increasingly polluted, especially urban ones. Is it possible to increase self
purification capacity as a tool for water recovery? The objective of this article was to test the increase
of biofilm and macrophythe management (BioMac) as a way of increasing the self purification
capacity in a polluted tropical urban river, using bioremediation as a river depollution tool.
Macrophytes management and artificial structures for biofilm agreggation were instaled in the river,
being a process of biorremediation rby bioestimulation with the provisiono of habitat. Physical,
biological and chemical parameters wer analysed. The results showed that after the biotreatment
insertion there was a great decrease of the nutriente concentrations along the river, allowing an
increase of the transparency, the biological diversity and the increase of the dissolved oxygen. This
effect was stronger in the most polluted site, Cabelo River headwaters, that receives sewers. It was
concluded that the BioMac pilot biotreatment system, was effective and of low cost, decreasing
phosphate compounds, increasing nitrification processes, the oxygen concentrations and the aquatic
plant diversity and the clarifying water, so it can be used in sewage rich rivers, such as in urban
environments.
Keywords: Biotechnology, Biotreatment, Periphyton, Macrophytes.
01

2. 1 – INTRODUCTION
The recovery of aquatic ecosystems is
now a demand and can be a problem, due to the
high cost of labor, products and machinery
invested. More accessible and low-cost
techniques, with methodologies that use
resources from the environment are welcome.
The Brazilian Ministry of the Environment-
MMA (2019) differentiates recovery from
restoration of degraded areas, emphasising that
the first is “restoring an ecosystem or a degraded
wild population to an undegraded condition,
which may be different from its original
condition”, while the second is the "restoration of
a degraded ecosystem or wild population as close
as possible to its original condition". As these
terms are used for actions that induce lasting
changes, the term depollution will be used in this
research.
There are several ways to restore the
aquatic environment, one of which is through
bioremediation. According to Gaylarde et al.
(2005), bioremediation is an ecologically more
appropriate and effective alternative for the
treatment of environments contaminated with
organic molecules of difficult degradation and
toxic metals. Currently it has also been used to
reduce the existing pollution in aquatic bodies,
since, according to the National Water Agency-
ANA (2017), only 39% of the organic load is
removed from the more than nine thousand tons
of sewage generated daily in Brasil, index well
below the 60% minimum removal set by the
National Environment Council - CONAMA.
Several studies have emphasized the
bioremediation carried out by microorganisms
(Vymazal, 1988, Crispim et al., 2009; Wu et al.,
2014; Ni et al, 2018; Ma et al., 2018), however,
macrophytes can also be used efficiently in the
reduction of nitrogen and phosphate compounds
(Biudes e Camargo, 2012; Crispim et al., 2019)
and as it serves as food for fish and other animals,
it encourages environmental recolonization by
biota. Wu (2017) comments that Periphyton is an
essential part of healthy ecosystems and can
retain nutrients and chemicals by various
processes. One of the bioremediation
methodologies that can be used is to increase the
area of occupation of the biofilm, using artificial
substrates, in order to promote the growth and
increase of the biomass of this community
(Crispim et al., 2009; Lu et al., 2014; Pérez,
2015; Ma et al., 2018; Morashashi et al, 2019;
Crispim et al, 2019;), thus increasing the self-
purification capacity of aquatic ecosystems.
Research carried out by Marinho (2018)
emphasizes an increase in the diversity of fish
species when increasing the biofilm in the
aquatic environment together with the whole
environment improve.
Thus, the hypothesis tested in this
research is “It is possible to improve water
quality in a polluted river, using biofilm and
macrophytes management, method we named
“BioMac”, as a depollution tool” and the aim of
this work is to test the effect of bioremediation,
through the use of biofilm and macrophyte
management, in the depollution of Cabelo River,
Paraíba - Brasil, being this research pioneering in
lotic environment with the present methodology.
2 - MATERIAL AND METHODS
Study Area
The experiment was realized in situ in a
tropical river. The Cabelo River is located in the
João Pessoa city among the coordinates
7°08’53’’ and 7°11’02’’ South and 34°47’26’’
and 34°East at an average altitude of 31.15m.
With an extension of 6.5 Km flows into the
Atlantic Ocean and border the beaches Penha
(south) and Seixas (north) (Fig 1). It is important
to mention that in dry period, the water of the
Cabelo River source doesn’t flow and the river
water is the Prison Institution sewer dumped,
forming a lake, and probably some percolation
water from subterranean waters to this lake,
following the river to P1.
The sampling points selection was related
to impacts and differences in local
characteristics. P1 is located 50 m downstream
from the sewage release; P2 is located after the
river passes through an area with natural
characteristics, although it sometimes receives
sewage discharges from a sewage transport
elevator; P3 is located after the river passes
through a remnant of the Atlantic Forest and
receives contributions from another spring that
also receives sewage; P4 is located in a partially

3. dammed area with the presence of many
macrophytes; P5 is located after a large
macrophyte bank; P6 is located at the mouth of
the river and has marine interference with the
collections always carried out at the ebb tide.
This river is classified as Class 3 based in
357/2005 CONAMA Resolution. In all sampling
sites, except P6, biotreatment was inserted.
Before P1, in the prison settling pond, biofilm
modules and macrophytes were installed.
Nevertheless, in this site the macrophytes did not
survive long. In P2 e P3, due the low depth,
stones and construction waste was used, getting
below the water level. In P4, P5 both biofilme
modules and macrophytes were installed.
FIGURE 1: Cabelo River in Paraíba State (red line), showing the source (yellow circle), the 6 collections sites (stars)
and their mouth in Atlantic Ocean (red arrow).
Font: Tiago Gusmão, 2022

4. Monitoring
The present study is a long-term study,
along 4 years, and was divided into 2 periods.
Monitoring without Biotreatment –BioMac
(period before inserting the bioremediation
structures from June/15 to November/16) and
monitoring with biotreatment (period after the
insertion of the biotreatment, from September
2017 to February 2018). In late January, the
bioremediation structures were removed, to see
their influence on water quality.
Limnological analysis were monitored
monthly and analyzed Temperature, Electric
Conductivity, pH, Dissolved Oxygen,
Nitrogenous and Phosphates compounds. For
this, the following methods were used:
nitrogenous compounds and phosphates
following APHA (Clesceri et al., 1998), pH
(pHmeter - Hanna), conductivity
(conductivimeter - Technal) and dissolved
oxygen and temperature through an oximeter
(Hanna), analyzed in the field.
All samples were analyzed/collected in
ebb tide, to have less sea interference.
The present work builded bioremediation
structures for the biofilm aggregation (in P1,P4
and P5) and also managed macrophytes,
removing the excess, preventing them from
covering the entire surface of the water, as well
as were used stones and left over building
materials (broken bricks) found in the
environment itself, to favor colonization by
biofilm (P2 and P3) where the river has low
depth, monitoring and analyzing the results with
the aim of checking the interference of BioMac
in the water quality of the Cabelo River.
Although the macrophytes have been used as
phytoremediators the use of the set of biofilms
and macrophytes (BioMac) is a ground breaking
project, being tested in a river for the first time
(Fig 2).
Macrophytes were retained inside a
floating circle, with 1 m diameter, nevertheless,
the macrophytes in the river grew too much and
control was lost in some stretches, and then they
had to be removed manually.
.
FIGURE 2: Containment structure for the macrophytes (with 1 m diameter) for the bio treatment BioMac, installed in
the Cabelo River in September 2017.
Substrate structures for the biofilm
aggregation – Several substrate modules for
biofilm agreggation were built using crystal
polyethylene (0. 30 mm). They were putted like
curtains in water column, with nylon and 2 L pet
bottles for floating. Empty pet bottles of 200 mL
were filled with sand and trapped at the end of
the curtain in order for maintain the curtain in a
vertical position. The square structure was
adapted with nylon (Fig 3) and measured 1,5m
each side and 1,0 m deep. (Fig 4-C), having 5
curtain rows inside each module, being equally
separated with knots for not to join (Fig 4-B).
In each corner 2 empty plastic bottles
were positioned for floatation and in each side for
mooring with knots two more bottles were
inserted separately (Fig 4 – A) On the two side
without mooring and in each of the inside rows
(which hold the plastic curtain) 1 plastic bottle
was inserted in the central part of the rows for
vertical position maintenance of the curtain.

5. Above the water level only the pet bottles
can be seen (Fig 4 -D)
FIGURE 3: Scheme of the bioremediation structures seen from above (left) vertical arrangement of 5 plastic curtains
under the water (right and above) and details of each plastic showing the eyelets, the wire and the 200 ml bottle (right and
below).
Statistical analysis
were carried out comparing the periods
without (before) and with Biotreatment (After),
for each sampling site. Tests for normality were
done using Shapiro-Wilk. When the data were
parametric test-T was applied for the non
parametric data Wilcoxon test was applied. The
cluster test was used to classify the elements into
groups, and the variables used for the test were
nitrite, nitrate, total phosphorus, orthophosphate
and ammonia. All the test was used through the
program R-Studio (2018).
Self-cleaning
To verify the self-purifications
capacity, the diference between the
concentrations of nutrientes and oxygen from P1
to P6 was calculated as a percentage value,
according to the equation below (simple rule of
3). As it was found that from P4 onwards some
nutrientes increased again, it was decided to also
estimate the depuration capacity in two more
stretches, between P1 and P4 and between P4 and
P6. In this case, in the stretch between P4 and P6,
the concentrations recorded in P4 and no longer
in P1 were considered as the 100% value.
Eq. 01
Eq. 2 X% – 100%
MS – Average parameter data in the stretch start
point (P1, P4) (without and with biotreatment)
ME – Average parameter data in stretch end
point (P6, P4)
X – Percentual of the parameter in the end
streatch (P4 ou P6)
100% the parameter value in the source streatch
(P1 or P4).
MS = 100%
ME = X%

6. FIGURE 4: Bioremediation structures showing the nylon and the bottles in the corner (A) the predefined moorings of
the structure (B) the structure finished showing the arrangement of the plastic curtains (C) and the structure in the river
(D).
3 - RESULTS
In the period without biotreatment the
largest precipitations happened on May/16
(166.4 mm) and June/15 (107.3 mm) whilst the
lowest was on December/15 (0.5 mm) and
February/16 (0,0 mm). In the period with
biotreatment the largest precipitations occurred
on February/18 (138.1 mm) and the lowest on
December/17 (0.5 mm) (Fig 5). Taking into
consideration that between October/17 and
January/18 were the periods with the lowest
precipitation rates, the positive results shown in
this research suffered very little interference
from the dilution caused by rainwater so this
result was mainly due to the biotreatment with
BioMac. Despite the fact that from February
onwards the bioremediation modules will no
longer be available, it can be seen later (Results)
that even with the rain, the tendency of the water
quality was to deteriorate again, revealing the
relevance of the biofilm increase.
The lower water temperatures
registered in the majority of the months (06/15,
07/15, 10/15, 11/15, 12/15, 02/16, 03/16 and
05/16) between June/15 and November/16
were in P3, whilst the higher were in P6. As
from September/17 on the lowest temperature
occurred on February/18 in P5 (26.7o
C) and
the highest was 32ºC (March/18) in P3 (Fig. 6-
above).
The statistical analyzes carried out
between the sites before and after the
installation of the biotreatment showed
significant differences in temperature in P1, P2
e P3 (Fig 6 -below), being lower in P1 and
higher in P2 and P3, respectively.

7. FIGURE 5: Accumulated monthly precipitation (mm) in the João Pessoa city in the period from Jun/15 to Mar/ 18.
Font: AESA – Precipitation (2019)
FIGURE 6: Monthy variation of air and water temperatures (ºC) (above) and Significant differences (below) between
the sampling sites without and with biotreatment in the Cabelo River.
With regard to the pH variation, without
biotreatment, the highest value was registered in
P3 on March/16, when achieved 8,8 and the
lowest was in P6 on December/15 with 4.6,
although in the majority of the months the
highest values were registered in P1. With
biotreatment, the lowest value was registered on
January/18 in the Lagoon (4.9) and the highest
pH value was 7.6 on January/18 in P2, lower than
before biotreatment (Fig. 7). Statistical analyzes
carried out between each site before and after the
installation of biotreatment, did not show any
significant differences to this parameter.
Among the study sites from P1 to P5, P1
registered the highest conductivity values in the
majority of the months in both periods, not
beyond 950 µS.cm-1
. The P6 conductivity values
were not considered in the statistical analysis,
due the presence of mangrove. In this site (P6)
conductivity varied from 236.2 µS.cm-1
in
January/18 to 906.8 µS.cm-1
. in October/15. The
lower values were registered in P5 in the majority

8. of the months, followed by P2, having this last
the lowest value registered in May/16 with 11.3
uScm-1
(Fig. 8 –above) The statistical analysis
comparing before and after biotreatment, carried
out for P1 (n=12, t=2.6596, p=0.0449), P4 (n=12,
t=3.9109, p=0.0113), P5 (n=12, t=4.0465,
p=0.0098) and P6 (n=12, v=21, p=0.03125)
showed significant differences, with lower
values after the installation of BioMac, whilst in
P2 and P3 weren’t significant (Fig 8-below)
FIGURE 7: Monthy variation of pH between the sampling sites without and with biotreatment in the Cabelo River.
FIGURE 8: Monthy variation of Electrical Conductivity (µS/cm) (above) and Significant Differences (below) between
the sampling sites without and with biotreatment in the Cabelo River.

9. With regard to the dissolved oxygen
concentrations, without biotreatment, the higher
values were registered in P6 in the majority of the
months reaching 8.70 mg.L-1
on March/16,
whilst the lower concentrations were in P1 in the
majority of the months, however, the lowest
concentration was registered in P4 (0.04 mg.L-1
)
on September/16. During the period with
biotreatment, the higher values was registered in
P2 (7. 4 mg.L-1
in September/17 and 8.6 mg.L-1
in January/18), P3 (6.5 mg.L-1
in September/17
and 6.1 mg.L-1
in November/17) and in P6 (7.1
mg.L-1
in November/17 and 6.7 mg.L-1
in
September/17). The highest value was 8.60
mg.L-1
in P2 (Fig. 9-above). The statistical
analysis comparing each sampling site before
and after the insertion of BioMac, showed
significant differences in P1 and P2 (increasing
oxygenation) and in P4 and P6 (decreasing
oxygenation) (Fig 9-below).
Total phosphorus concentrations without
biotreatment obtained higher values registered in
P1 in all the months, reaching a maximum
concentration of 3.65mg.L-1
on October/15. With
biotreatment, the highest concentrations
registered continued being in P1 (Fig. 10-
above).The statistical analysis carried out with
P1 (n=24, v=262, p< 0,001), P3(n=24, v=219,
p=0.0490) and P6 (n=24, v=283, p=2.468e-05
),
comparing both periods showed significant
differences, showing lower concentrations after
biotreatment (Fig. 10-below), whilst P2, P4, and
P5 showed no significant diferences.
FIGURE 9: Monthy variation of Dissolved Oxygen (mg.L-1
) (above) and Significant differences (below) between the
sampling sites without and with biotreatment in the Cabelo River.

10. FIGURE 10: Monthy variation of Total Phosphorus concentration (mg.L-1
) (above) and Significant differences (below)
between the sampling sites without and with biotreatment in the Cabelo River.
The highest concentrations of
orthophosphate were registered in P1 as well,
concentrations being usually above 2.0 mg.L-1
.
With biotreatment even though P1 continued
showing the highest values, these tended to be
lower than in the period before the biotreatment,
not being higher than 2.2 mgL-1
(Fig. 11-above).
The statistical analysis carried out showed P3
(n=24, v=221, p=0.012) and P4 (n=24, v=189,
p=0.011) having significant differences
comparing before and after the BioMac
treatment, with lower concentrations after the
implantation of the biotreatment, however for P1,
P2, P5 and P6 there were no significant
differences (Fig. 11-below).
To ammonia a similar pattern to
phosphorus, was observed, with larger
concentrations in the first sites of samples, in the
region of the main source, but with significantly
lower concentrations after the insertion of
BioMac. In the period without biotreatment, the
largest concentration was 5.32 mg.L-1
(April/16)
in P1, whilst with biotreatment, it was on
November/17 with 3,30mg.L-1
in P1 (Fig 12-
above). The statistical analysis comparing both
analyzed periods carried out with P1 (n=48,
v=256, p=0.003), P4 (n=48, v=284.5 p=0.001),
P5 (n=48, v=300, p=1.192e07
), and P6 (n=48,
v=300, p=1.192e-07
) showed significant
differences, with lower concentrations after
biotreatment whilst P2 and P3 did not show any
significant differences (Fig 12-below).

11. FIGURE 11: Monthy variation of Orthophosphate concentration (mg.L-1
) (above) and Significant differences (below)
between the sampling sites without and with biotreatment in the Cabelo River.
FIGURE 12: Monthy variation of Ammonia concentration (mg.L-1
) (above) and Significant differences (below) between
the sampling sites without and with biotreatment in the Cabelo River.

12. The higher concentrations of nitrite were
registered in P2 and P3 in the most months,
during the whole study period, with a maximum
value of 0.5 mg.L-1
in P2 (January/18) in the
period with biotreatment, coinciding with the
lack of rainfall in that month (Fig. 13-above).
The statistical analysis carried out in P1 (n=48,
v=52.5, p=0.029) and P3 (n=48, p=0.019)
showed significant differences, reaching the
highest nitrite concentrations after the
biotreatment, whilst in P2, P4, P5 and P6 they
were not significant (Fig 13-below).
The higher values of nitrate were
registered in P3 in most months in the period
without biotreatment, however, the highest
concentration was 5.66 mg.L-1
in P2 (July/15). In
the months with biotreatment, the higher values
were registered in P2 in the majority of the
months with a highest concentration of 4.4 mg.L-
1
on December/17 (Fig. 14-above). The statistical
analysis carried out for P1 (n=28, v=4.5
p<0.001), P2 (n=28, v=52, p=0.009), P3 (n=28,
v=14, p=1.311e-05
) showed significant
differences, with higher values after the
installation of BioMac whilst in P4, P5 and P6 no
differences could be found (Fig. 14-below)
FIGURE 13: Monthy variation of Nitrite concentration (mg.L-1
) (above) and Significant differences (below) between
the sampling sites without and with biotreatment in the Cabelo River.

13. FIGURE 14: Monthy variation of Nitrate concentration (mg.L-1
) (above) and Significant differences (below) between
the sampling sites without and with biotreatment in the Cabelo River.
Analysing the self depuration capacity in
Cabelo River, from source to the mouth, it was
observed a great increase in depuration after the
insertion of the biotreatment, from 54% to 90,6%
for example in the ammonium concentration, 0%
to 90% in nitrite concentrations, etc. The oxygen
concentration depuration capacity also changed,
in some stretches (Table I). However, it appears
that the depuration capacity is not the same in all
the analyzed stretches of the river.
For example, in relation to oxygen
concentrations, there is a depuration capacity
between P1 and P6 before biotreatment, reaching
an increase of this gas of about 440%, due to the
low concentrations that were registered in P1, it
was verified increase in oxygen along the river.
After biotreatment, this percentage reduced to
about 100%. On the one hand, because due to
biotreatment, oxygen concentrations improved in
P1, but on the other hand, because in the P4
region, there were many floating macrophytes,
which, due to their decomposition, reduce the
oxygen concentration, due to the increase of
decomposing microorganisms. When analyzing
between P1 and P4, there is a reduction in
oxygen, which did not occur before. With better
water quality, more plants grew in this region,
reducing light input, which reduces
phytoplankton production with the respective
release of oxygen to the water on the one hand
and increased decomposition on the other. With
the reduction of oxygen in the stretch P1-P4,
there is an increase of this gas between P4 and P6
by 174%.
In relation to phosphate compounds,
changes were also observed between the two
periods, before and after biotreatment. While
before bioremediation there was a decrease in
orthophosphate concentrations from the
beginning to the end of the river, after
biotreatment, it appears that the entire reduction
of this compound occurred between P1 and P4,
with no change from this to P6 . The presence of
large amounts of macrophytes, despite absorbing
phosphate, also release it through decomposition,
keeping this balance in balance, without reducing
it. On the other hand, total phosphorus showed
similarity between P1 and P4 in the two periods,
but increased the purification capacity in the final
stretch, between P4 and P6, from 66.6% to
82.4%.

14. Table I: Average Self purification capacity of Cabelo River presented in % of decrease or increase (-) of limnological
parameters, in the periods without biotreatment (set/15 a fev/16) and with biotreatment (set/17 a fev/18).*data of
parameter = 0,0 in start and end of the stretch. **0,0 in start, not allowing the division.
Nutrient/site
WITHOUT
BIOTREATMENT
WITH BIOTREATMENT
P1-P6 P1-P4 P4-P6 P1-P6 P1-P4 P4-P6
Ammonia % 57,3 54 7,1 90,6 68,6 70,1
Nitrite % 100,0 100,0 * 90 100 **
Ortophosfate% 99,5 98,1 71,1 99,2 99,2 0,0
Total Phosphorus % 97,4 92,1 66,6 98,3 90,7 82,4
Oxigen - 441,5 - 171,9 - 99,2 - 102,3 26,1 - 173,7
Font: Draw up by author
Analysing the river environmental
conditions, before and after the biotreatment,
through a cluster analysis (phosporus,
orthophosphate, ammonia, nitrite and nitrate), is
possible to see that all sample station before (A),
excepting P1, grouped, and all sampling stations
after de biotreatment (D), excepting P6, grouped
as well, showing the clear differences on
environmental conditions before and after
applied the biotreatment (Fig 15). P1, before
biotreatment performed another group, due the
worst environmental conditions at all, but after
de biotreatment grouped with the otther sampling
stations. P6 also grouped alone after the
biotreatment because it showed clear better
conditions. This allowed the mangrove to born
and grow in the river mouth.
Through visual analysis, there were
differences in the dynamics of the Cabelo River
in P5 in relation to macrophyte diversity which
increased in the period with treatment, becoming
more biodiverse and changing dominant species,
which was no longer Pistia stratiotes and became
initially Nymphaea sp. and then Marsilea sp., and
at the end of 10 months, in P6, the early
colonization of mangrove species at the Cabelo
River mouth, on the beach was registered (Fig
16).
FIGURE 15: Cluster analysis comparing environmental
parameters before (A) and after (D) the biotreatment
(BioMac) application.
It is important to state that visually it was
observed the difference in the macrophytes
dominance in P5, before and after the insertion of
BioMac, with the larger diversity (increased from
5 species to 8) in the periods with Biotreatment,
and changing the dominance from Pistia
stratiotes to Nymphaea sp and then to Marsilea
sp., as well as the beginning of a colonization of
mangrove plant species in P6, showing a plant
regeneration process in the river mouth, changing
a species indicator of eutrophication as P.
stratiotes (Thomaz and Bini 2003) for Marsilea
sp. Nevertheless, this species increased great
banks, over all surface and it was necessary to
manage them taking out some of them, as a
management tool.
FIGURE 16: Cabelo River at P5 during the period without treatment with the presence of macrophyte Pistia stratiotes
dominating during the period of drought (A1) and during the rainy season (A2), during the biotreatment period in P5 (B1)
and in P6 (B2), with the development of mangrove. 2015 (A1 e A2) e 2017 (B1 e B2)

15. Photos: Flávia Oliveira (A1, A2 e B1) and Cristina Crispim (B2).
FIGURE 17: Management of macrophytes carried out monthly in P5 of Cabelo River, being removed manually (A and
B), with a sieve (C), broom (D) and being transported in the car bucked (E).

16. Photos: Flávia Oliveira.
4 – DISCUSSION
The main source of pollution in the
Cabelo River is found in the main source (before
P1), which in general makes the ecological
dynamics and biota of the entire downstream
river worse. P1 which reached the highest values
of pH, higher concentrations of total phosphorus,

17. ortophoshate and ammonia was due to the sewer
coming from the prisional Institutions nearby,
rich in grey and black waters, with the presence
of soap which liberate OH radicals (Esteves,
1998). This sampling site was also the one who
showed the highest electrical conductivity levels,
(with the exception of P6, in the estuary) due to
the presence of salts in the water also justified by
the sewers. Tundisi (2003) commented that good
fresh water quality is fundamental for the
sustainability of nutrient cycles, economic
development, and a better quality of life for the
population. The lower values of OD and elevated
concentrations of nutrients, these being,
ammonia, orthophosphate, and total phosphorus,
were higher at the sampling site of largest
anthropic action (P1), which receives largest
amount of discharge of organic matter (sewage
from the prison and from the elevatory sewage
system of CAGEPA), a fact also confirmed by
Sardinha et al. (2008) to this river.
It is important to note that the water
temperatures at sampling points 1, 2 and 3
diverged unproportionally from the air
temperatures (meaning lower air temperature and
higher water temperature) in some months,
mainly in the period without biotreatment, while
in the period with biotreatment this divergence
was milder, following it in some months.
Sampling points 4, 5 and 6 followed air
temperatures in most months, in both periods,
thus showing how greater pollution interferes
with water temperature, since P 1 was
characterized by being the most polluted, while
points 2 and 3 receive sewage discharge a few
times and interference from clandestine sewage
in the second source. As can be seen, the water
temperature was generally higher than the air
temperature, but was influenced by it. According
to Esteves (1998), polluted waters, which have
more particulate matter, have higher
temperatures, because these particles absorb
heat, transferring it to the water.
P4 was where the lowest values of pH
were obtained in the majority of the months that
were analyzed, and according to the values
established by CONAMA Resolution 357/2005,
for Class 3 river, must be between 6,0 and 9,0.
The pH found was some times below these limits
in some sampling sites, reaching 2,9. The largest
frequency of the lowest values associated with P4
is because in this place there are large
macrophytes banks, whose decomposition
induces a decrease in pH due to the release of
CO2, in parallel, lower oxygen concentrations
were registered in this area as well, due the
increase in decomposing microorganisms,
especially on nov/16. Pérez (2015) in a study in
this river, comparing different sizes of
macrophytes banks, in larger banks, oxygen
concentrations and pH were both lower,
comparing with the same streaches under smaller
macrophytes banks. Where decomposition
occurs CO2 is liberated, promoting acidity in the
water (Esteves, 2011). In the decomposition of
organic matter, fungi and heterotrophic bacteria
are of fundamental importance, as detritivorous
biota like Protozoa, sometimes making aquatic
ecosystems poor or absent in oxygen and often
causing death of aerobic aquatic organisms. Von
Sperling (1996) commentated that decomposing
bacteria are the group with the largest presence
and importance in increasing DBO values.
Riverside population reported episodes of
fish mortality in some periods, coinciding with
the opening of the seawage transport system
directly to the river, due to problems in the
pumping station, demonstrating that only in these
sporadic situations, the Cabelo River became
unsuitable for the aerobic biota to the point of
killing it.
The ammonia concentrations were higher
in P1 all the study period, nearby 3,0 (+0,5)
mg.L-1
, especially before de biotreatment, due
the presence of a sewage stabilization pond built
to treat sewage from prisons, which was placed
in the bed of the Cabelo River. The insufficient
treatment of this type of sewage treatment plant,
is responsible for all the loss of water quality
along the river, although the data show that the
river is sparsely inhabited until the mouth, and it
manages to improve its water quality.
Nevertheless, even maintaining the stabilization
pond, the biotreatment was effective,
significantly decreasing the ammonia
concentrations to values nearby
The highest values of nitrite and nitrate
were achieved in P2 and P3 in the period without
treatment, as a consequence of the nitrification
processes which occurred along the river as a
degradation process from ammonia. However, as
in P1 there was little oxygen, nitrification poorly
occurred. As in P2 the oxygen increases, making
possible the transformation of ammonia to nitrite,

18. these concentrations increased. The increase in
oxygen, favors nitrification (Dajoz 2005) and
inhibits denitrification (Zielinska et al 2012)
because denitrifying bacteria contain enzymes
that are inactivated in the presence of dissolved
oxygen (Zoppas et al. 2016). However, after
biotreatment these concentrations of nitrite and
nitrate even got higher, due the higher oxygen
concentrations that were present, improving
nitrification processes. Despite the plants
absorbing nitrate, as the amount of ammonia
present in the environment was high, due to the
presence of raw sewage, with the increase in
oxygenation, the ammonia passed more
efficiently to nitrate, maintaining an excess of
this in the water, compared to what was being
consumed by plants.
In P3, was registered a slight increase in
ammonia. This is the result of the increase in
pollution at the font of a second source of the
Cabelo River before P3, which begun to receive
effluents from the sewer of a gated community of
the region. The sites P1 and P3 showed
significant differences in the concentrations of
nitrite, with the highest concentration with
biotreatment, due to the better oxygenation
condition which favored the nitrification
dynamics, as told above. This increase in nitrite
concentrations was also observed by Lima
(2019) after using biotreatment, that induces
increase in oxygen concentrations.
Comparing the conductivity values
before and after the installation of BioMac, there
was a significant reduction in these values,
especially in P1, the nearest to the contamination
site, showing the effect of BioMac in the lake
which receives the sewer. This was seen along
the river, reducing the conductivity values at
other sampling sites as well, specially between
Sep/17 and Fev/18, showing an increase in the
self-healing capacity promoted by the
biotreatment (BioMac), fact proved by the
statistical analysis which showed significant
differences in the P1, P4, P5 and P6 after the
biotreatment, to this parameter.
In relation to Total Phosphorus, a positive
effect was registered by the insertion of BioMac
in all the monitored sampling sites. P1 continued
as being the place with the highest concentration
of Total Phosphorus, however, showed lower
values than the previous period in drought period
(September to March) changing the maximum
values from 3.65 mg.L-1
(October/15) to 2,86
mg.L-1
(October/17), and the other sampling sites
showed a large decrease in the concentration of
this compound, especially from November on,
showing clearly a decrease in eutrophication.
Wetzel (1996) commented that the
periphyton plays an important role in the
renovation tax of nutrients in the environment
and in accordance with Jones et al. (2002), the
periphyton can compete with the macrophytes
for carbon and light, just as interfering in the
transfer of nutrients between the pelagic and
benthic zones. This nutritional dynamic can
affect the colonization time. In the presente study
it was observed nearly 15 days for the
colonization of a substrate by the biofilm,
agreeing to Borduqui (2011) that registered
exponential growth until 12th day by periphyton.
In Orthophosphate concentrations, it was
found that the maximum concentrations
decreased with BioMac treatment, as well as the
concentration of this component along the river,
even without the rains (October/18 until
January/18) proving the efficiency of the
bioremediation system, reducing, for example
from 2.3mg.L-1
(September/15) and 2.4mg.L-1
(September/16) to 0.41mg.L-1
(September/17) as
well as from 2.9 mg.L-1
(November/15 and
November/16) to 2.6mg.L-1
(November/17) and
from 1.6. mg.L-1
(February/16) to 0.7mg.L-1
(February/18) all in P1. Ni et al, (2018) verified
that the biofilm that formed on the biofilter
played a major role in removal of organic
pollutants and nitrogen.
It can be said that the biotreatment by
bioremediation, with the use of biofilm,
increased the purification capacity of the Cabelo
River, mainly with the reduction of ammonia,
which is a toxic compound, and total phosphorus.
Oxygen, despite the depuration capacity having
been lower after biotreatment, as the increase in
the biofilm P1 started to present higher
concentrations of this gas, the environment
started to present itself more oxygenated.
However, in P4, there was a reduction of this gas,
due to the increase of macrophytes, which
affected the downstream purification capacity.
The reduction of ammonia concentrations
to values below 1mg.L-1 favored the appearance
of mangroves at the mouth of the river, which
was not observed before.

19. The application of BioMac in the Cabelo
River, apart from improving the water quality,
increasing the diversity of macrophytes it also
increased the number of fish species present
which increased from 6 to 15 (Marinho 2018)
which analyzed the ichthyofauna assembly
before and after the insertion of BioMac in this
river. The presence of 15 fish species, was the
same number of fish species in a less impacted
river nearby (Marinho 2018) (based in the used
capture methodology by the author) being
probably a number present in healthy mangroves
in this region. Because polyethylene is among the
polluting materials of aquatic environments and
generate residuals, like nanoplastic, based on a
research, previously carried out by Pérez (2015),
that also used plastic curtains to increase biofilme
in a dam, the author observed that it began to
deteriorate after 8 months of installation, than it
is suggested that the polyethylene curtains should
be substituted every 6 months, as it is a lotic
environment and as such exerts more pressure on
the plastics.
However, research into the use of more
ecological materials is necessary which Aquatic
Ecology Laboratory (Labea/UFPB) is already
carrying out by testing other materials, like
natural loofah (Luffa cylindrical) and sisal
(Agave sisalana) rope, which are derived from
plants, which apart from being a non-pollutant,
can be removed and used as animal food or
organic compost, together with biofilm.
5 – CONCLUSIONS
It can be concluded that the pilot system
of biotreatment in rivers with BioMac was
efficient and of low cost, and can be used in
rivers which receive large quantities of sewage,
as in Brazilian urban environments. In general, a
great influence was seen of biotreatment with
BioMac, with a significant decrease in the
concentrations of nutrients or an increase in
oxygen and transparency (this last one, visual
observation), when the environmental conditions
were worst as in P1, P4, and P5, with the
exception of nitrogenous compounds nitrite and
nitrate, which showed a significant increase in
the most oxygenated sites (P2 and P3) permitting
the transformation of ammonia into these
compounds, due the increase in oxygen
concentrations.
The period of one month was the time to
check the differences in the surroundings, a
period necessary for the installation of biofilm in
the substrate, showing that the use of
polyethylene was adequate for the creation of
new habitats for biofilm, increasing the self
purification capacity of the river, leading to an
water quality improve in Cabelo River. In
informal conversation with resident people and
direct observation a decrease in the bad smell
was also felt. Visually it was possible to observe
an increase in the transparency, an increase in
fish fingerlings in P4, a return to fishing in P5 and
P6, the emergence of mangrove plants in P6 in
response to the depollution of the river.
Thus, the BioMac depollution system is
presented as an efficient and low-cost way to
clean up polluted urban rivers, mainly those
polluted by untreated sewages.
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______________________________________
1. Doutora pelo Programa de Pós-graduação em
Desenvolvimento e Meio Ambiente
(PRODEMA); Centro de Ciências Exatas e da
Natureza da Universidade Federal da Paraíba
(UFPB). Orcid: https://orcid.org/0000-0002-
6974-138X ; E-mail: fmf_oliveira@hotmail.com
2. Profa. Dra. do Departamento de Sistemática e
Ecologia/Centro de Ciências Exatas e da
Natureza/ Universidade Federal da Paraíba.
Cidade Universitária; CEP 58059-900, João
Pessoa, PB, Brasil ORCID:
https://orcid.org/0000-0002-4414-2989;
21