Biofouling

The Journal of Bioadhesion and Biofilm Research

ISSN: (Print) (Online) Journal homepage: https://www.tandfonline.com/loi/gbif20

Identification of bacterial biofilms on desalination

reverse osmosis membranes from the
mediterranean sea

Zakaria Benladghem , Sidi Mohamed Lahbib Seddiki & Yassine Moustafa

Mahdad

To cite this article: Zakaria Benladghem , Sidi Mohamed Lahbib Seddiki & Yassine Moustafa
Mahdad (2020): Identification of bacterial biofilms on desalination reverse osmosis membranes
from the mediterranean sea, Biofouling, DOI: 10.1080/08927014.2020.1851366

To link to this article: https://doi.org/10.1080/08927014.2020.1851366

Published online: 24 Nov 2020.

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BIOFOULING
https://doi.org/10.1080/08927014.2020.1851366

Identification of bacterial biofilms on desalination reverse osmosis

membranes from the mediterranean sea
Zakaria Benladghema , Sidi Mohamed Lahbib Seddikia,b and Yassine Moustafa Mahdadb,c
a
LAPSAB Lab: Antifungal Antibiotic, Physico-Chemical Synthesis and Biological Activity, University of Tlemcen, Tlemcen,
Algeria; bDepartment of Biology, University Center of Na^ama, Na^ama, Algeria; cPPABIONUT Lab: Physiology,
Physiopathology and Biochemistry of Nutrition, University of Tlemcen, Tlemcen, Algeria

ABSTRACT ARTICLE HISTORY

Nanofiltration and reverse osmosis are two of the most effective surface water treatment Received 24 May 2020
processes. They provide water of high quality and eliminate a large amount of microorganisms, Accepted 10 November 2020
organic matter and micropollutants. However, the main limitation of membrane nanofiltration is
KEYWORDS
fouling, which imposes an additional cost. This study focused on the search for microorganisms
autoaggregation; bacterial
capable of reducing the performance of nanofilters and also to study autoaggregation and biofilms; biofouling;
biofilms formation by bacterial strains isolated from the nanomembranes used in the seawater nanofiltration;
desalination plant of Souk Tlata (Algeria). It provides new microbiological data on the reverse osmosis
desalination of seawater in the southern Mediterranean basin. The results revealed 14 bacterial
species isolated from six fouled reverse osmosis membranes; their quantities were significant
with the dominance of Raoultella sp., Klebsiella sp., Staphylococcus sp., Stenotrophomonas sp.,
Micrococcus sp., and Escherichia coli. In addition, electron imaging of nanomembrane surfaces
revealed complex structures of microorganisms forming biofilms.

Introduction filtration technique used in the desalination of sea-

Water is essential to maintain life on earth; it is
a highly strategic resource on which many human
activities depend. In natural environments, water can
contain many chemicals and/or biological contamin-
ation which can have serious repercussions on the
environment and the health of consumers in the case
of drinking water (Levi et al. 2007). On the other
hand, the sea is the largest water reserve on the
planet, but it remains unfit for consumption due to
its high content of electrolytes, organic materials,
microorganisms, algae and certain pollutants such
as petroleum and toxic industrial waste (Kettab 2001;
Litumanya et al. 2018). Like other Mediterranean
countries, Algeria is faced with the problem of
decreasing water reserves due to the scarcity of rain
as well as its irregular frequency and, already based
on the mobilization of natural resources by capture
(e.g. dams, reservoirs), reflecting a painful and uncer-
tain water policy (Kehal 2001; Shannon et al. 2008).
However, this country has 1,200 km of coastline,
which gives it the alternative of the desalination
of seawater (Kettab 2001). This process uses the
principle of reverse osmosis (RO), a membrane
water, brackish water and also in the reuse of urban
and industrial wastewater. This filtration process
removes the salts and organic substances present in
the water such as bacteria and viruses (Gaid and
Treal 2007). However, the main disadvantage of
membrane nanofiltration is fouling (Van den Broek
et al. 2010), including particle fouling, inorganic (scal-
ing) and organic fouling and biofouling. Biofouling is
one of the most serious forms of fouling of mem-
branes affecting the performance of the process of
reverse osmosis (Al-Juboori and Yusaf 2012; Nguyen
et al. 2012; Khan et al. 2013). Although some biofilms
are beneficial for environmental detoxification in
bioreactor technology, purification of waste water and
waste air, for soil remediation and the decomposition
of solid waste (Flemming 1993), other biofilms cause
biofouling of RO membranes. They are difficult to
control or prevent by existing cleaning procedures in
which the microorganisms, like members of the
Gammaproteobacteria, adhere on the surface of the
nanomembranes, form biofilms and secrete extracellu-
lar polymeric substances (EPS) which protect them
against chemical cleaning agents (Flemming et al.

CONTACT Zakaria Benladghem benladghemz@gmail.com

ß 2020 Informa UK Limited, trading as Taylor & Francis Group
2 Z. BENLADGHEM ET AL.
1997; Vrouwenvelder and Van der Kooij 2001; Di
Martino et al. 2007; Belila et al. 2016). In addition,
the structure of biofilms formed on different areas of
the surface of RO membranes varies depending on
external factors such as shear force, salt concentration,
pressure changes and permeate flow (Nagaraj et al.
2017). According to the available literature, no study
has been carried out in this context on the southern
Mediterranean coasts, in particular those of Algeria.
The aim of this study was to investigate the develop-
ment of biofilms on the surface of the seawater reverse
osmosis (SWRO) membranes used in the desalination
plant of Souk Tlata (Algeria). Additional objectives
included searching for Gammaproteobacteria and
Gram-positive bacteria on the surfaces of fouled
SWRO membranes, to assess their potential to form
biofilms as well as their capacity to autoagregate, to
determine the relationship between these two phenom-
ena and finally, to demonstrate the three-dimensional
structure of biofilms by scanning electron microscopy.
Material and methods
Membrane sampling
The sampling was carried out between September
2018 and September 2019 in the Souk Tlata seawater
desalination unit which is made up of thirteen
identical SWRO filtration trains for a production
capacity of 200,000 m3 day21. SWC5 and SWC5 MAX
(Hydranautics, Nitto Group Company) were the
membranes used; they are made of polyamide and
have a minimum salt rejection rate of 99.7%.
Eighteen samples were taken from six fouled RO
membranes. From each membrane, previously placed
in a laminar flow hood, three fragments, each
measuring 1 cm2, were removed and placed in tubes
containing 10 ml of sterile nutrient broth (Sigma-
Aldrich, Germany). Samples were taken from different
locations of the membrane; one at the entrance, one
in the middle and one at the exit.
Isolation and biochemical tests
After incubation of the samples in nutrient broth for
24 h at 37
C, the strains were successively inoculated
on liquid and solid culture media as well as on Mac
Conkey and Chapman (Sigma-Aldrich, Germany),
respectively. 37
C is a favorable temperature for the
culture of marine bacteria. Since the water in the
desalination plants is intended for human consump-
tion it is important to incubate at this temperature
which is favorable to the culture of pathogenic
bacteria (Priyanka Das et al. 2019).The dishes were
thereby incubated again at 37
C for 24 to 48 h.
Galleries API 20E, API 20NE and API Staph
(Biomerieux, La Balme les-Grottes, France) were used to
identify Gammaproteobacteria, Staphylococcus spp. and
Micrococcus spp. among the strains isolated. The API
system is a miniaturized version of conventional
biochemical techniques; a bacterial suspension equiva-
lent to 0.5 Mc Farland was distributed in the wells, the
galleries were then incubated at 37
C for 24 h. The
strains were identified by referring to the identification
table of the analytical catalog as well as to the following
software (API 20E þ V 4.1, API 20NE V 7.0 and API
Staph V 4.1). All tests were performed in duplicate.
Biofilm formation potential
The technique of staining with crystal violet makes it
possible to measure the quantity of biomass in the
biofilm (Christensen et al. 1985). 20 ml of pre-cultures
of each isolated bacterial strain were incubated for
24 h at 37
C. The bacterial suspensions were centri-
fuged (1,000 g, 15 min, 4
C), and then the pellets
were washed with sterile phosphate buffered saline
(PBS, 10 mM, pH 7.4); this operation was performed
twice. Bacterial cells were then resuspended in Tryptic
Soy Broth culture medium (TSB, pH 7.3), chosen
to form the biofilms. Inocula concentrations were
fixed at 107 cells ml1 (0.5 Mc Farland), then 100 ll
of each were introduced into the wells of a sterile
96-well microplate. The plates were sealed and then
incubated at 37
C for 24 h.
After biofilm formation, the medium was aspirated
and the wells were washed three times with sterile
PBS to eliminate planktonic cells and/or non-adherent
cells. For fixation of the biofilm, 100 ll of methanol
(99%) were added to each well and then incubated
for 15 min at room temperature. The wells were
washed and filled with 100 ml of a crystal violet solu-
tion (0.1% w/v). The plates were incubated again for
20 min at room temperature before being washed
once more. The crystal violet linked to the biofilms
was released by the addition of 150 ll of acetic acid
(33%) to each well, the absorbance was then revealed
at 570 nm using a microplate reader (Rayto, RT-
2100C) indicatinging the quantity of biofilm biomass.
The cut-off OD (ODc) for the microplate test was
defined as three standard deviations above the mean
OD of the negative control. According to (Stepanovic
et al. 2000), the potential to form biofilm of the strains
tested was classified into the following categories:
 BIOFOULING 3

Table 1. Strains isolated from six fouled SWRO membranes in the desalination plant at Souk T’lata-Algeria.
Isolated strains Number (%) Gram tests Percentage accuracy (%)
Escherichia coli 7 (19) 2 95-99
Raoultella ornithinolytica 3 (8) 2 81-89
Raoultella terrigena 1 (2.7) 2 70
Klebsiella oxytoca 4 (11) 2 94-97
Klebsiella pneumoniae 1 (2.7) 2 98
Enterobacter amnigenus 1 (2.7) 2 97
Serratia odorifera 1 (2.7) 2 99
Stenotrophomonas maltophilia 3 (8) 2 99-100
Pseudomonas luteola 1 (2.7) 2 98
Micrococcus spp 5 (13) 1 97-99
Staphylococcus xylosus 4 (11 ) 1 52-77
Staphylococcus cohnii ssp cohnii 3 (8) 1 95-99
Staphylococcus haemolyticus 1 (2.7) 1 97
Staphylococcus saprophyticus 1 (2.7) 1 64
Staphylococcus sciuri 1 (2.7) 1 56
Total 37 (100) / /

ODc< OD  2  ODc weakly adherent; 2  ODc
< OD  4  ODc moderately adherent;
4  ODc<OD strongly adherent. All tests were
performed in duplicate.
Autoaggregation assay
Autoaggregation was analyzed using two methods.
Firstly, using visual test in tubes (Rupani et al. 2008),
which was revealed by the formation of aggregates at
the bottom of the tube and the clarity of the suspen-
sion during incubation. Secondly, by the quantitative
spectrophotometric method (Karched et al. 2015)
with some modifications; the ability of the isolated
strains to form cellular aggregates was quantitatively
tested by measuring the absorbance at 600 nm.
Briefly, cell suspension grown for 24 h in nutrient
broth for each strain was centrifuged (5,000 g, 15 min,
4
C) and washed twice with KCl buffer (pH 6.0,
50 mM KCl, 1 mM CaCl2, 1 mM KH2PO4, 0.1 mM
MgCl2). The turbidity was adjusted to 1.0 at 600 nm
and an aliquot of 1 ml of the solution was pipetted
into a microcuvette. The measurement of the percent-
age autoaggregation was carried out by the decline in
the optical densities of the bacterial suspensions after
incubation for 24 h. To that, the OD was measured
immediately at 600 nm (time zero value) and after
24 h (sample value). The autoaggregation rate was cal-
culated by the following equation:
% Autoaggregation ¼ ½time zero value
sample value=time zero value  100Þ
Scanning electron microscopy
Scanning electron microscopy (SEM) was used to
highlight details of biofilms that formed on the sur-
face of the fouled SWRO membrane. The samples
intended for SEM observation were taken from the
outlet areas of fouled SWRO membranes, and then
placed in sterile PBS.
Before their examination, the nanomembranes were
rinsed with PBS, and then fixed with a glutaraldehyde
solution (2.5% v/v). The samples were observed under
the ZEISS EVO 10 electron microscope.
Statistical analysis
The comparison between strain means was studied
using analysis of variance (ANOVA) and Duncan’s
Multiple Range Test (p < 0.01) using GenStat
Discovery Edition 3 statistical software.
Results
Isolation and biochemical tests
From six fouled SWRO membrane surfaces, a total of
37 bacterial strains were isolated and purified. Among
them, 22 (59%) were classified as Gram-negative and
15 (41%) as Gram- positive (Table 1).
Gram-negative bacteria belonged to the class of
Gammaproteobacteria. These were 18 (82%) strains
belonging to the family Enterobacteriaceae, and 4
(18%) non-fermenting bacteria including three
Stenotrophomonas maltophilia and one Pseudomonas
luteola. For Gram-positive bacteria (Table 1),
Micrococcus spp. and Staphylococcus spp.
were identified.
Regarding the number of species isolated from the
different areas of fouled SWRO membranes, the outlet
contained the most bacterial strains (43%), followed
by the middle zone (32%) and finally by that of the
inlet (25%). Enterobacteria and some Gram-positive
bacteria appeared to be the predominant species that
colonized the clogged surfaces of the
SWRO membranes.
4 Z. BENLADGHEM ET AL.

Figure 1. Quantitative results of biofilms formed by isolated strains based on the crystal violet method. Bars with the same letters
do not differ significantly from Duncan’s Multiple Range Test at the 1% probability level. Vertical bars show standard errors (SEs).
ROi1-A - ROi36-N: isolated strains.

Biofilm formation and autoaggregation

It should be noted that the ability to form biofilms
and autoaggregation tests were carried out by
repeated absorbance measurements. Fourteen bacterial
strains were selected from each species to perform
these tests; the strains that showed the best ability to
form biofilms were chosen.
According to the results obtained, 11 strains (78%)
were classified as good biofilm formers (Fig. 1).
Pseudomonas luteola (ROi33-k) had the greatest sig-
nificant potential to form biofilm (p < 0.01) with a
mean OD570 of 2.25 ± 0.02. This was followed by
Stenotrophomonas maltophilia (ROi15-E), Klebsiella
oxytoca (ROi31-I) and then Raoultella ornithinolytica
(ROi32-J), respectively with a mean OD570 of
1.62 ± 0.01, 1.60 ± 0.02 and 1.60 ± 0.01; the difference
between these results was not significant with regard Figure 2. Macroscopic observation of the autoaggregation test
to the biofilm formation potential of these three after incubation of the strains ROi33-k (A) and ROi34-L (B) for 24 h.
strains (Fig. 1). The following strains were, respect-
ively, listed in descending order; Staphylococcus sciuri Pseudomonas luteola (ROi33-k) was the strongest
(ROi35-M), Staphylococcus haemolyticus (ROi16-F), autoaggregating strain with 64 ± 2% followed by
Raoultella terrigena (ROi3-C), Micrococcus spp. Staphylococcus haemolyticus (ROi16-F) then
(ROi36-N), Enterobacter amnigenus (ROi9-D), Staphylococcus xylosus (ROi28-H), respectively with
Klebsiella pneumoniae (ROi2-B), and Staphylococcus 57.76 ± 3.24% and 57.46 ± 1.54% (Fig. 3). For these
cohnii (ROi17-G). On the other hand, Serratia odori- three strains, no significant difference was recorded
fera (ROi34-L) and Escherichia coli (ROi1-A) were with regard to autoaggregation; however, the results
classified as significantly moderately biofilm-forming were significant compared with the rest of the strains
(p < 0.01). Likewise, Staphylococcus xylosus (ROi28-H) (p < 0.01). On the other hand, Serratia odorifera
was weakly biofilm-forming (p < 0.01). (ROi34-L) had the weakest capacity to form cellular
For the autoaggregation test, all strains were able aggregates with a rate of 26.13 ± 1.13%. This result was
to form cellular aggregates. Overall, macroscopic ana- not significantly different from that of Micrococcus spp.
lysis of autoaggregation test revealed bacterial cells (ROi36-N), Klebsiella oxytoca (ROi31-I) and
which aggregated and settled at the bottom of the cul- Stenotrophomonas maltophilia (ROi15-E).
ture tube under static incubation (Fig. 2A). While the The results obtained revealed that Pseudomonas
tubes of strains with low autoaggregation potential luteola (ROi33-k) presented a dual high potential to
remained cloudy (Fig. 2B). form a biofilm and cellular autoaggregation (Fig. 4). On
 BIOFOULING 5

Figure 3. Autoaggregation potential of strains isolated from obstructed SWRO membranes. Bars with the same letters do
not differ significantly from Duncan’s Multiple Range Test at the 1% probability level. Vertical bars show SEs. ROi1-A - ROi36-N:
isolated strains.

SWRO membrane; deposits, in particular crystals,

were observed. But at higher magnification (Fig. 5B),
the structure of the mixed biofilms was evident; many
bacteria spread irregularly across the surface of the
membrane. However, the presence of single cells and
cells forming compact aggregates was revealed.
Different bacterial morphologies were observed on
the surface of the membrane, viz. cocci, bacilli and
bacteria in the shape of large rods.

Discussion
Figure 4. Scatter plot of the isolated strains as a function of
the autoaggregation rate and the biofilm formation potential. This study demonstrates the alteration of the SWRO
membranes used in the Souk- tlata desalination plant
(Algeria). The results obtained showed that these
the contrary, Serratia odorifera (ROi34-L) had the lowest
alterations were caused by abiotic and biotic substan-
potential for biofilm formation and also for autoaggrega-
ces, bacteria in particular. Indeed, biofouling in water
tion. However, Staphylococcus xylosus (ROi28-H) and
treatment processes is the major reason for poor plant
Escherichia coli (ROi1-A) which showed the lowest rate
performance (Chiellini et al. 2012). Among all the
of biofilm formation had, conversely, a high potential to
strains isolated, the dominance of Gram-negative
form cell aggregates; these potentials were however
bacteria was notable. This corresponds to the results
reversed in Stenotrophomonas maltophilia (ROi15-E).
of Ivnitsky et al. (2007) who showed that the bacterial
Taking into account these results, the in vitro tests
species isolated from the nanofiltration membranes
for biofilm formation and autoaggregation showed no
applied to wastewater treatment were mainly
correlation between them since the correlation coeffi-
Gram-negative. According to the results obtained, 14
cient was 0.065 and the P value was 0.744. This suggests
bacterial species were identified; it is however very
that these two phenomena are independent of each
difficult to determine the full range of micro-
other, despite the fact that the results obtained for the
organisms in seawater (Nagaraj et al. 2017). This
strains ROi33-K and ROi34-L seem to have, respect-
study showed that the majority of the Gram-negative
ively, a positive and negative correlation between them.
bacteria isolated belonged to the class
Gammaproteobacteria; while Dang et al. (2008) point
Scanning electron microscopy
out that this class of bacteria has been shown to be
The surface of the fouled RO membrane was covered among the most dominant bacterial assemblage in
by a mixture of bacteria and abiotic deposit. coastal marine ecosystems. In this context, Nagaraj
Figure 5A shows a complex heterogeneous structure et al. (2017) found that Gammaproteobacteria
which uniformly covered the entire surface of the dominated the bacterial community in moderately
6 Z. BENLADGHEM ET AL.
Figure 5. SEM images of (A): a mixture of deposits substances and (B): microbial biofilms on the surface of a fouled SWRO
membrane. Arrows indicate bacterial cells with different morphology. The bars ¼ 20 mm [magnification2,000] (A) and 2 mm
[magnification 8,000] (B).
fouled SWRO membranes. On the other hand, Belila
et al. (2016) found that Gammaproteobacteria were
the second colonizer among six classes of
Proteobacteria isolated from SWRO membranes.
According to H€ orsch et al. (2005), this group of bac-
teria is the main colonizer of nanofiltration membranes
on which they form biofilms. In addition, Bereschenko
et al. (2007, 2008) showed that the feed water to the
RO system, permeate from cartridge ultrafiltration,
contained a wide variety of phylotypes typical of pure
water, among them Gammaproteobacteria. According
to this study, 7 strains of E. coli were isolated with a
percentage accuracy of 95 to 99%, which represented
19% of all the Enterobacteria isolated. Indeed, seawater
has been presented as a relevant reservoir of E. coli
strains that have adapted to the marine environment
(Alves et al. 2014). In this context, a recent study
(Nagaraj et al. 2017) pointed out that a high abun-
dance of Enterobacteria, including Escherichia spp.,
was recorded in samples of cartridge filters. For Gram-
positive bacteria, the results of this study show that
27% of the strains isolated were Staphylococcus species.
Previous research has indeed found several species of
Staphylococcus in marine samples (Fang et al. 2006;
Harakeh et al. 2006). In addition, the same studies
have shown that the high level of contamination by
staphylococci in most samples makes it possible to ver-
ify the ability of this microorganism to survive in diffi-
cult environmental conditions.
Micrococcus isolates in this study represented
13%. This result shows that this species can survive
in marine environments. Nevertheless, several studies
have shown that the sea is a rich reservoir of
Micrococcus, which is especially found in seawater,
sediments and marine sponges (Wood 1952;
Mohapatra and Bapuji 1997; Umadevi and
Krishnaveni 2013).
The bacterial community of biofilms formed on
the surface of SWRO membranes represents a very
small proportion of the bacterial community con-
tained in seawater (Nagaraj et al. 2017). Moreover,
the conditions on the surface of nanomembranes are
selective for microbial populations and, consequently,
the cells embedded in the biofilms and the EPS layer
become more difficult to remove over time (de Vries
et al. 2019).
The present study showed different levels of bio-
film formation in all bacterial isolates. Previous work
has shown that phylotypes of Gammaproteobacteria
were numerically abundant in the SWRO membrane
biofilms community (Pang and Liu 2007). Moreover,
Bereschenko et al. (2010) have shown that bacteria of
this class can detach from mature biofilm and adhere
as aggregates to other surfaces of the SWRO mem-
brane and therefore develop other microcolonies. This
suggests that the outlet of the SWRO membrane is
the most contaminated part compared with other
areas. It seems that the biofilms at the exit ends of
the SWRO membrane were adapted to the different
oligotrophic conditions compared with the inputs and
the middle regions; this may be due to a lower per-
meate flow caused by the resistance of the fouling
layer when the feed water circulates along the mem-
brane. On the other hand, Stenotrophomonas malto-
philia, with a percentage accuracy of 99 to 100%,
represents  8% of the strains isolated from SWRO
membranes; previous research had established that
the Stenotrophomonas phylotype represented  10%
of the biofilm community formed on these mem-
branes (Pang and Liu 2007). In addition, according to
the results obtained in this study, Pseudomonas spp.
had a high potential to form biofilms; a previous
study noted that this bacterium was classified as a
good biofilm former on SWRO membranes (Belgini

et al. 2014). It was also one of the main agents
responsible for the biological fouling of SWRO mem-
branes in seawater desalination plants (Belila et al.
2016). In addition, Al Ashhab et al. (2017) found that
Pseudomonadaceae were among the most dominant
families in all RO membranes samples. Overall, mar-
ine biofilms hide significant microbial diversity
(Zhang et al. 2019). It has been found that bacterial
strains isolated from the Mediterranean Sea exhibit
different abilities to form biofilm (Brian-Jaisson et al.
2014); this result parallels that of the present study.
The results of the autoaggregation tests revealed all
the strains isolated were able to form cellular aggregates.
But Pseudomonas luteola had the highest autoaggrega-
tion potential and Serratia odorifera had the lowest.
Autoaggregation is a phenomenon widely observed in
environmental species. It is among the first steps in the
construction of a biofilm and it can lead to the forma-
tion of microcolonies and therefore to a biofilm (Trunk
et al. 2018). In this study, there was no correlation
between the potential for biofilm formation and autoag-
gregation in any of the bacterial isolates. Effectively,
according to Trunk et al. (2018), this phenomenon does
not always favor the formation of biofilms. This is not
in agreement with others (Sorroche et al. 2012) which
have revealed a positive correlation between bacterial
autoaggregation and biofilm formation.
On the other hand, high resolution SEM images
were used to observe the microorganisms present on
the membrane, possibly responsible for biological foul-
ing at the time of their removal. The images showed
complex structures of abiotic deposits and bacterial
cells adhering to the surface of the SWRO membranes.
These results are in agreement with those found by
Bereschenko et al. (2010) who showed that the mature
biofilm formed on SWRO membranes displayed a
complex heterogeneous structure and was propagated
over the entire membrane surface. However, protobio-
films which are transparent exopolymer particles, are
free-floating microgel particles which participate in bac-
terial colonization and the formation of aquatic bio-
films (Berman and Holenberg 2005; Bar-Zeev et al.
2012; Nagaraj et al. 2018). Further, compared to bio-
films that are attached to surfaces, protobiofilms float
freely (Bar-Zeev et al. 2012).
Conclusions
Biofouling of the nanomembranes used in the filtra-
tion of seawater is a problem in the desalination pro-
cess. The main objectives of the current study were to
search for Gammaproteobacteria and Gram-positive
BIOFOULING 7
bacteria on the surfaces of fouled SWRO membranes
and to determine the relationship between the poten-
tial for biofilm formation and the autoaggregation
capacity of the isolated strains, and also, to reveal the
three-dimensional structure of the biofilms formed on
the membrane surfaces. This study provides new
microbiological data on the desalination of seawater
in the southern Mediterranean basin (Algeria). It also
reinforces previous data from other studies carried
out in different maritime sites.
Gram-negative and Gram-positive bacteria were
the main colonizers of these membranes and the
results revealed the deposit of Gram-negative and
Gram-positive bacteria on the surfaces of the SWRO
membranes. Enterobacteria were the most representa-
tive, notably E. coli, Raoultella spp. and Klebsiella spp.
For Gram-positive bacteria, Micrococcus spp. and
Staphylococcus spp. were in the majority. Indeed, elec-
tronic imaging has shown the deposition of abiotic
substances and a mixture of bacterial cells forming
biofilms; this leads to biological fouling. Tests for the
potential for biofilm formation and autoaggregation,
carried out in vitro, revealed a variable capacity in the
strains tested; however, there was no correlation
between these two biological phenomena. Mature bio-
films formed on the surface of SWRO membranes are
difficult to detach or control, therefore a solution
must be found to reduce the microbial concentration
in the water before it reaches these membranes. In
order to develop better control strategies, it is essen-
tial to broaden the field of future research, not only
to bacteria but also to viruses, algae and protozoa.
Acknowledgements
The authors wish to thank Ahmed Dayri, director of the
Souk T’lata seawater desalination station, and Ismail
Guermat, technical engineer, for their help and assistance.
Disclosure statement
The authors declare no conflict of interest.
ORCID
Zakaria Benladghem http://orcid.org/0000-0001-
9929-3779
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