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Cultivating microalgae in wastewater for biomass production, pollutant remov-

al, and atmospheric carbon mitigation; a review

Ayesha Shahid, Sana Malik, Hui Zhu, Jianren Xu, Muhammad Zohaib Nawaz,
Shahid Nawaz, Md. Asraful Alam, Muhammad Aamer Mehmood

PII: S0048-9697(19)35295-7
DOI: https://doi.org/10.1016/j.scitotenv.2019.135303
Reference: STOTEN 135303

To appear in: Science of the Total Environment

Received Date: 7 August 2019

Revised Date: 27 October 2019
Accepted Date: 29 October 2019

Please cite this article as: A. Shahid, S. Malik, H. Zhu, J. Xu, M.Z. Nawaz, S. Nawaz, Md. Asraful Alam, M.A.
Mehmood, Cultivating microalgae in wastewater for biomass production, pollutant removal, and atmospheric carbon
mitigation; a review, Science of the Total Environment (2019), doi: https://doi.org/10.1016/j.scitotenv.2019.135303

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 Cultivating microalgae in wastewater for biomass production, pollutant removal,
and atmospheric carbon mitigation; a review

Ayesha Shahid 2, Sana Malik 2, Hui Zhu 1, Jianren Xu 3, Muhammad Zohaib Nawaz 4,5, Shahid
Nawaz 6, Md. Asraful Alam 7*, Muhammad Aamer Mehmood 1,2*

1 School of Bioengineering, Sichuan University of Science and Engineering, Zigong 643000,

People’s Republic of China

2 Bioenergy Research Centre, Department of Bioinformatics & Biotechnology, Government

College University Faisalabad, Faisalabad 38000, Pakistan

3 College of Bioscience and Engineering, North Minzu University, Yinchuan 750021, Ningxia,

China

4 State Key Laboratory of Marine Environmental Science, Institute of Marine Microbes and

Ecospheres, College of Ocean and Earth Sciences, Xiamen University, Xiamen, China

5 Department of Computer Science, The University of Agriculture Faisalabad, Faisalabad 38000,

Pakistan

6 Department of Chemistry, The University of Agriculture Faisalabad, Faisalabad 38000,

Pakistan

7 School of Chemical Engineering, Zhengzhou University, Zhengzhou 450001, China

Corresponding author’s email: draamer@gcuf.edu.pk (MA Mehmood, Ph.D.), alam@zzu.edu.cn

(MA Alam, Ph.D.)

Authors note: First two authors contributed equally to this paper and should be considered as co-
first authors

1
 Cultivating microalgae in wastewater for biomass production, pollutant removal,
and atmospheric carbon mitigation; a review

Ayesha Shahid 2, Sana Malik 2, Hui Zhu 1, Jianren Xu 3, Muhammad Zohaib Nawaz 4,5, Shahid
Nawaz 6, Md. Asraful Alam 7*, Muhammad Aamer Mehmood 1,2*

1 School of Bioengineering, Sichuan University of Science and Engineering, Zigong 643000,

People’s Republic of China

2 Bioenergy Research Centre, Department of Bioinformatics & Biotechnology, Government

College University Faisalabad, Faisalabad 38000, Pakistan

3 College of Bioscience and Engineering, North Minzu University, Yinchuan 750021, Ningxia,

China

4 State Key Laboratory of Marine Environmental Science, Institute of Marine Microbes and

Ecospheres, College of Ocean and Earth Sciences, Xiamen University, Xiamen, China

5 Department of Computer Science, The University of Agriculture Faisalabad, Faisalabad 38000,

Pakistan

6 Department of Chemistry, The University of Agriculture Faisalabad, Faisalabad 38000,

Pakistan

7 School of Chemical Engineering, Zhengzhou University, Zhengzhou 450001, China

Corresponding author’s email: draamer@gcuf.edu.pk (MA Mehmood, Ph.D.), alam@zzu.edu.cn

(MA Alam, Ph.D.)

Authors note: First two authors contributed equally to this paper and should be considered as co-
first authors

2
Abstract
The water shortage is one of the leading global problems along with the depletion of
energy resources and environmental deterioration. Recent industrialization, global mobility, and
increasing population have adversely affected the freshwater resources. The wastewater sources
are categorized as domestic, agricultural and industrial effluents and their disposal into water
bodies poses a harmful impact on human and animal health due to the presence of higher
amounts of nitrogen, phosphorus, sulfur, heavy metals and other organic/inorganic pollutants.
Several conventional treatment methods have been employed, but none of those can be termed as
a universal method due to their high cost, less efficiency, and non-environment friendly nature.
Alternatively, wastewater treatment using microalgae (phycoremediation) offers several
advantages over chemical-based treatment methods. Microalgae cultivation using wastewater
offers the highest atmospheric carbon fixation rate (1.83 kg CO2/kg of biomass) and fastest
biomass productivity (40-50% higher than terrestrial crops) among all terrestrial bio-remediators
with concomitant pollutant removal (80-100%). Moreover, the algal biomass may contain high-
value metabolites including omega-3-fatty acids, pigments, amino acids, and high sugar content.
Hence, after extraction of high-value compounds, residual biomass can be either directly
converted to energy through thermochemical transformation or can be used to produce biofuels
through biological fermentation or transesterification. This review highlights the recent advances
in microalgal biotechnology to establish a biorefinery approach to treat wastewater using
microalgae. The articulation of wastewater treatment facilities with microalgal biorefinery, the
use of microalgal consortia, the possible merits, and demerits of phycoremediation are also
discussed. The impact of wastewater-derived nutrient stress and its exploitation to modify the
algal metabolite content in view of future concerns of cost-benefit ratios of algal biorefineries is
also highlighted.

Keywords: microalgae cultivation; wastewater treatment; biorefineries; CO2 fixation; cost-

effective
1. Introduction
Dawn of the 21st century led the world towards increasing urbanization, industrialization
and commercialization at the cost of increasing carbon emission, depleting energy and water
resources, and contamination of water bodies with toxic pollutants (Khan et al., 2019). It is
expected that the world’s population will increase from 6.3 billion in 2015 to approximately 9
billion in 2050 (Ng et al., 2017). The increased population would require enough and cleaner
energy supply as well as the clean water resources. To acquire and maintain a clean environment,
it would be needed to reduce CO2 content by 50-80% with an increase of 50% in water and
energy resources respectively (Hightower and Pierce, 2008; Ng et al., 2017). According to 2014
statistics, estimated global freshwater consumption was 3700 billion m3 (Diniz et al., 2017),
where most of it is being converted into wastewater due to human activities. Based on its origin,
wastewater produced by human activities is termed as domestic, aqua-cultural, agricultural or
industrial sewage/effluents (Diniz et al., 2017). Various industries in paper and pulp industry,
sugarcane industry, textile or tannery industry, and pharmaceutical industry (Lv et al., 2017;
Ramlow et al., 2017) contribute to surface water pollution causing water scarcity. Polluted and
untreated water contaminates freshwater resources by releasing excess amounts of nitrogen and
phosphorus which enhances eutrophication and ecosystem destruction, hence making it
unsuitable for human consumption (Godfray et al., 2010; Morée et al., 2013). Water pollution
(acidification, eutrophication, sewage, heavy metals and other organic pollutants) (Godfray et al.,
2010) have confirmed harmful effects on human health, including growth inhibition,
feminization of male organisms, carcinogenicity, (Butkovskyi et al., 2017) development of
waterborne diseases like diarrhea, typhoid, intestinal worms, gastroenteritis, Cryptosporidium
infections, cardiovascular, renal failure and hypertension (Daud et al., 2017), both in developed
and developing countries.

The main aim of wastewater treatment is to remove the excess amount of micropollutants
(Grandclement et al., 2017), nutrients (nitrogen, sulfur, copper, phosphorus) (Wang et al.,
2017a), heavy metals (copper, zinc, lead, mercury, chromium, nickel, cadmium) (Bilal et al.,
2013) and organic pollutants (phenolic compounds, aromatic hydrocarbons, biocides, surfactants,
antibiotics etc.) from wastewater (Salama et al., 2017). A variety of chemicals, physical and
biological wastewater treatment methods have been employed (Wang et al., 2017a) while the
biological approach being more common (Grandclement et al., 2017) to remove these pollutants.
4
Depending on the effluent requirement, the wastewater treatment method is selected (Dvořák et
al., 2014). However, there is no standalone method that can be applied to a variety of effluents
because of the limitations associated with each method (Table 1). In general, conventional
treatment methods have shortcomings such as the requirement of large land, intensive energy
input, and extensive maintenance and operational costs (Udaiyappan et al., 2017). Hence, it is
required to develop alternative technologies to recycle wastewater along with the fixation of
atmospheric carbon (Hariz and Takriff, 2017). Microalgae-based bioremediation is the safest,
promising, and the most efficient alternative replacing conventional treatment methods due to its
vast availability, higher nutrient consumption ability and diverse applications of the algal
biomass produced (Lam et al., 2012).

This review summarizes different wastewater treatment methods and focused on cost-
effective and efficient microalgae-based of wastewater (phycoremediation) along with the
potential problems and opportunities (Fig. 1). It also highlights the possibilities of exploiting the
wastewater as a low-cost growth media and as a natural stress-manipulative for enhanced
biomass productivity and algal metabolite content, presenting an algal-biorefinery concept.

2. Microalgae-based wastewater treatment

Algae are one of the most diverse groups of eukaryotes starting from simple blue-green
algae to complex sea-weeds and kelps. On average, more than 350,000 microalgal species have
been discovered (Shahid et al., 2017). In general, they are believed to contain as high as 70%
lipids, 60% carbohydrates and 65% of proteins and essential amino-acids respectively (Afzal et
al., 2017). Microalgal biomass is an alternative of traditional feedstocks (Khan et al., 2018)
because microalgae have a short growth cycle when compared to terrestrial plants or energy
crops, higher biomass productivity, higher harvesting index, and the highest rate of carbon
fixation. Additionally, microalgae don’t require large arable land instead can be grown on
marginal lands by using seawater or wastewater as growth media (Miranda et al., 2017).
Microalgae play a vital role in environmental carbon mitigation and bioremediation due to their
higher photosynthetic efficiency (40-50% higher than terrestrial plants) (Chen et al., 2015), CO2
sequestration (1 kg of microalgae consumes 1.83 kg of CO2 and accounts for 40% of global CO2
sequestration) (Chisti, 2007; Ng et al., 2017). Microalgae can also be used as bioindicators to
detect the climate changes in aquatic environments (O'Neill et al., 2019) and can consume
wastewater nutrients (80-100% uptake of nitrogen and phosphorus) for high productivities of
biomass and value-added products (Grandclement et al., 2017; Miranda et al., 2017; Su et al.,
2016). Depending upon the microalgal strains, they can be employed in various industries like
cosmetics, poultry, biofertilizers, medicine and green-fuels namely bioalcohols, biogas and
biodiesel (Afzal et al., 2017). However, it is required to develop cost-effective strategies to
produce cost-competitive algal products, because at present the algal biofuels cannot compete the
prices of the fossil fuels (Dasan et al., 2019). Hence selection of appropriate strain (Gill et al.,
2016), use of low cost media, optimization of conditions for higher biomass production, cell
stoichiometry to divert the balance towards target product, suitable commercialization, and
reducing the operational cost (mainly associated with cultivation and harvesting stages) are
focused aspects of the algal research (Ng et al., 2017). Figure 2 summarizes a roadmap of
microalgal growth optimization and strain development for microalgal biorefinery.

Wastewater is the most suitable resource for algal biomass production because of several
reasons such as; (i) cheaper growth media, (ii) support bulk biomass and biofuel production, (iii)
can supply ample nutrients, and (iv) offers the possibility of integrating algal cultivation with the
existing infrastructure wastewater treatment (Roostaei and Zhang, 2016). Several studies have
been conducted in last decades on microalgae-based phycoremediation of wastewater (Chen et
al., 2015; Salama et al., 2017; Udaiyappan et al., 2017; Wang et al., 2017a) and biorefinery based
approaches have been proposed (Gill et al., 2013). Microalgae have been studied for wastewater
(industrial and domestic) treatment including brewery wastewater (Ferreira et al., 2017),
domestic wastewater (Calicioglu and Demirer, 2017), textile wastewater (Wu et al., 2017a),
pharmaceutical waste streams (Xie et al., 2019), slaughterhouse industry (Aziz et al., 2019),
heavy metal-containing wastewater (Khan et al., 2017), palm oil mill effluents (Hariz and
Takriff, 2017), starch-containing textile wastewater (Lin et al., 2017), and agro-industrial
wastewater (Jayakumar et al., 2017). Though microalgae-based phycoremediation has numerous
benefits, however, there are several challenges that also need to be addressed.

Several studies have demonstrated species of Chlamydomonas, Chlorella and

Scenedesmus can be employed for efficient nutrient uptake (N, P) (Gao et al., 2016) and removal
of toxic pollutants and heavy metals (Khan et al., 2017; Matamoros et al., 2015) from
wastewater. Ankistrodesmus along with Scenedesmus and Chlorella has shown to degrade

6
organic pollutants present in paper and oil mill industry wastewater (Bhattacharya et al., 2017).
Similarly, Scenedesmus sp. represented higher utilization efficiency of 98.2%, 97.1 % and
95.2% for butyrate, propionate, and acetate respectively with a growth rate of 0.53 gd-1 when
cultivated in textile desizing wastewater (Lin et al., 2017). Chlorella variabilis showed to
consume nutrients from textile effluents and with 100% remediation ability for nickel,
aluminum, and iron with biomass productivity of 74 gm-2d-1 with lipid yield of 20 %
(Bhattacharya et al., 2017). Similarly, high nitrogen (99.6%) and phosphorus (91.2%) removal
efficiency with the concomitant production of biomethane (523 mL) was reported by C. vulgaris
grown in municipal wastewater (Calicioglu and Demirer, 2017). Freshwater microalgae
including Cladophora glomerata and Oedogonium westii can remove heavy metals from
industrial wastewater and showed 80% of cadmium and 66 % of nickel accumulation
respectively (Khan et al., 2017). Additionally, Neochloris aquatica CL-M1 was employed to
produce butanol along with wastewater treatment (Wang et al., 2017c), where biobutanol yield of
0.89 g L-1 h-1 with 96.2% of NH3-N removal efficiency was observed. Moreover, lipid-rich
Botryococcus sp. removed 59.9% nitrogen, 36.8% phosphate and 54.5% organic carbon along
with the production of 72.5% of crude oil content, when cultivated in domestic wastewater (Gani
et al., 2017).

Biotic factors (microbial load including competing pathogens) and abiotic factors
(nutrients, pH, and CO2) play a pivotal role in microalgal metabolism. Manipulation of stress
factors including biotic and/or abiotic through wastewater significantly affects the productivities
of both biomass and product of interest (Table 2). Unraveling the underlying processes and
reorienting the related pathways can greatly contribute to achieving robustness, enhanced
productivities in energy and cost-efficient manner (Chen et al., 2017). Utilization of wastewater
as an alternative media offers additional benefits including wastewater cleaning, recycling,
reducing the environmental pollutions, and provision of low-cost growth media (Yadav et al.,
2019). However, it requires detailed studies of using wastewater as low-cost growth media and
studying the impact of biotic and abiotic factors to attain the full potential of microalgae for the
biorefinery.
2.1. Removal of macro and micro-nutrients

Microalgae store lipids as polar (polyunsaturated fatty acids) and non-polar forms
(saturated fatty acids, mainly in the form of triacylglycerides) which are important to maintain
membrane functions or to perform other cellular activities. These lipids can be transesterified to
produce biodiesel (Paliwal et al., 2017). The stress of various macro and micronutrients, mainly
nitrogen and phosphorus, has been studied widely to enhance algal metabolite content mainly
lipids.

2.1.1. Nitrogen

Nitrogen is one of the principal nutrients for the synthesis of nucleic acid, protein,
energy-carrying molecules (ATP), and enzymes (Juneja et al., 2013). Naturally, nitrogen is
available in the form of nitrate, urea, ammonium, and peptones (Minhas et al., 2016). In
microalgae, nitrogen accounts for 1-20% of dry cell matter and is part of the essential functional
and structural proteins of algae (Juneja et al., 2013). Nitrogen starvation has been considered as
the most effective induction strategy (Shi et al., 2017) that shifts the organism’s metabolic
pathway towards enhanced lipid synthesis, triglyceride accumulation, protein content reduction
and increased carotenogenesis, at the expense of biomass production (Juneja et al., 2013; Minhas
et al., 2016).

An increase of 2.27-fold in lipid content was observed in Scenedesmus quadricauda

when grown under nitrogen-starved conditions (226 mgL-1) (Anand and Arumugam, 2015).
Acutodesmus dimorphus accumulated 75% neutral lipids of total lipids under nitrogen-starvation
conditions (Chokshi et al., 2017). Cultivation of Pseudokirchneriella subcapitata under nitrate
stress conditions resulted in 160 mgL-1d-1 fatty acid accumulation (Del Río et al., 2017).
Scenedesmus obliquus showed 4.4 mgL-1day-1 of total nitrogen removal efficiency with 1.4 gL-1
of biomass and 29.8 mgL-1day-1 of lipid productivity when grown on urban wastewater (Álvarez-
Díaz et al., 2017). Similarly, removal of 84.51% of nitrate and 75.56% of ammonium along with
the production of 172.9 mgL-1day-1 of carbohydrate, 150.2 mgL-1day-1 of lipid and 141.5 mgL-
1day-1 of protein productivities were reported C. sorokiniana cultured in wastewater (Guldhe et
al., 2017). Cultivating C. vulgaris in mixed piggery and brewery wastewater reduced nitrogen
and ammonia by 96% and 100%, respectively producing 2.85 gL-1 biomass (Zheng et al., 2018).

8
Similarly, COD and nitrogen removal efficiencies of 76% and 98%, respectively, were reported
for wastewater treatment using S. obliquus (Gupta et al., 2016).

These studies have clearly indicated that nitrogen stress has a global impact on various
microalgae to enhance lipid content, but it cannot be applied on a commercial scale using
synthetic growth media, which will raise the cost. Alternatively, nitrogen stress can be applied
using wastewater, which often has higher nitrogen content (municipal wastewater), however,
wastewater from industrial sources may have lower nitrogen content. Consequently, water from
these sources can be used to exert nitrogen stress on the microalgae. Removal of nitrogen by
microalgae does not only treat wastewater but also trigger the accumulation of value-added
products. However, aore detailed studies are required in the future to elucidate the role of
wastewater derived N-stress on growth and biomass productivity of microalgae.

2.1.2. Phosphorus
Phosphate is another important (second only to nitrogen) macronutrient of a living system
which is part of RNA, DNA backbone, ATP, phospholipids (Juneja et al., 2013),
phosphoproteins, polyphosphates and in the form of NADPH. Phosphate limiting conditions
lead to organic carbon accumulation (TAG) and decreased cell division. Phosphorus deficiency
is also known to affect the energy-requiring processes like protein synthesis, transcription, and
carbon cycle (Mühlroth et al., 2017). Generally, it constitutes <1 to 1% of the total dry mass of
microalgae (Minhas et al., 2016). The C. protothecoides has shown to accumulate 32.8 % of lipid
content under mixotrophic conditions using wastewater as growth media which was deficient in
phosphorus (Li et al., 2014). Similarly, under nutrient stress, the C. vulgaris was shown to
accumulate lipids by 17.41% producing 9.81 gL-1 of biomass, which is comparatively low than
nitrogen and sulfur stress (Sakarika and Kornaros, 2017). Similarly, 53% of lipid content and
23.45 mgL-1 of lipid productivity was reported for Chlorella under phosphorus deprivation
conditions (Wong et al., 2017).

Phycoremediation of municipal wastewater by Micractinium sp. IC-76 resulted in 77% of

PO4-P removal efficiency with 37.18 mg L-1day-1 of biomass productivity and 36% of lipid
content (Piligaev et al., 2017). Cultivation of Desmodesmus abundans on synthetic wastewater
showed 16% of lipid content and 87.52% of phosphate removal rate (Prasad et al., 2017).
Treatment of sewage water by phosphate starved Scenedesmus (Yewalkar-Kulkarni et al., 2017)
resulted in an enhanced carbohydrate ratio with 87% of phosphate removal. Hence, it is
suggested to expose the microalgae to the combined stress of nitrogen and phosphorus to achieve
higher biomass and lipid productivities (Chen et al., 2017). A suitable cost-effective alternative is
the use of wastewater. Cultivation of microalgae in wastewater will consume phosphorus (act as
stress factor) and thus help in wastewater treatment.

2.1.3. Sulfur
Sulfur is also one of the most important macronutrients. Commonly, sulfate is consumed
as a sulfur source by plants and algae. Cellular processes namely assimilation, secondary
metabolic pathways, oxidative stress responses, flavonoid, and nitrogen metabolism are affected
by sulfur. In algae, enhanced assimilation of Sulfur is believed to downregulate the
photosynthesis, and its stress induces expression of stress-associated proteins (Giordano and
Raven, 2014). Naturally, sulfate is present in excessive amounts in the wastewater from paper
and pulp industry, pharmaceutical industry, mining, and food processing (Lv et al., 2017). An
increasing trend in neutral lipids (123-172%), oligosaccharide, TAG and polysaccharide levels
was observed for wild-type Chlorella strain cultivated under sulfur-deprived conditions (Cakmak
et al., 2012). Moreover, the sulfur starvation showed to increase the β-carotene level from 6.753
mgL-1 to 14.616 mgL-1 with decreasing biomass production in Dunaliella salina (Shaker et al.,
2017). Unlike Nitrogen, Sulfur starvation has proven to be more efficient for starch accumulation
when compared to Nitrogen starvation which causes lipid accumulation (Vitova et al., 2015). An
increase of 18-fold in starch content was observed in C. reinhardtii when cultured under S-
deprived conditions. It is believed that sulfur starvation inhibits energy-consuming pathways like
cell growth and division while, shifting the pathways towards starch accumulation (Antal et al.,
2014). Interestingly, Sulfur deprivation altered the fatty acid composition of C. lobophora
(Takeshita et al., 2014). On the other hand, sulfate stress negatively affected the pollutant
removal ability, growth and self-flocculation efficiency of Chlorococcum sp. GD when grown in
synthetic municipal wastewater (Lv et al., 2017). It indicates that the impact of sulfate may not
be global, hence we need to conduct carefully designed detailed studies to evaluate the potential
of pollutant removal, biomass and metabolite productivities under sulfur stress alone, and/or the
combined stress of other nutrients, which may lead towards the selection of suitable strains.
While the selection of strain will be influenced by the type and source of the wastewater too.

10
2.1.4. Heavy metals
Metals are the natural constituents of soil and earth crust. However, in ecological terms,
any metalloid or metal which has bioaccumulation ability and causes environmental pollution is
termed as heavy metal. Some of the metals are micronutrients (Cu, Zn, Ni, Mn, and Co) and are
essential for growth (Kumar et al., 2015), and some others like Mn, Cu, I, Zn, Fe, Pb, etc. are
proven to be beneficial for nutritional quality improvement and other important functions of
living system (Wells et al., 2017). Some heavy metals including Pb, Cd, Cr, and Hg have
unidentified biological functions and are toxic in nature (Afshan et al., 2015; Kumar et al., 2015)
as their excessive uptake may affect metabolic processes, the physical structure of algae and can
cause toxicity, mutagenesis and allergenicity (Mikulewicz et al., 2017).

Heavy metals stimulate the ROS (reactive oxygen species) formation resulting in
oxidative damage. Decreased metabolite content (pigments, proteins, and monosaccharides), size
and number of cells observed to be related to lead accumulation in Acutodesmus obliquus
(Piotrowska-Niczyporuk et al., 2015). Increased protein and carbohydrate ratios were observed
when C. sarokiniana cultivated in mixotrophic and photoautotrophic conditions under the
influence of titanium-dioxide (TiO2) nanoparticles (Marchello et al., 2018). Exposure of copper
sulfate (CuSO4) induced lipid peroxidation due to the formation of ROS and reduced the levels
of carotenoids, chlorophyll-a, and b in Chlorella, which reflects highly toxic nature of CuSO4
(Wan et al., 2018). Cheng and coworkers (Cheng et al., 2016) reported 71%, 93% and 74%
reduction in carotenoids, chlorophyll-a, and chlorophyll-b level, respectively, in C. vulgaris as a
result of 7 mgL-1 cadmium treatment while ~96% increase in soluble protein accumulation was
observed.

Modern agronomic practices (metal-based pesticides), intense industrialization (metal,

textile, mining etc.), enhanced anthropogenic activities (rubber, paint, paper, and metal alloys
production), and unauthorized waste disposal have increased the concentration of these
pollutants in environment (Das and Osborne, 2018; Kumar and Gunasundari, 2018). Biosorption
of heavy metals from wastewater by microalgae is a promising alternative as it is a cost-
effective, ecologically safe and efficient method (Ummalyma et al., 2018) when compared to
conventionally employed methods. C. vulgaris, Chlamydomonas reinhardtii, Chlorococcum spp,
Phaeodactylum tricornotum, Scenedesmus quadricauda, Spirogyra spp. and many other algal
species have been reported for heavy metal biosorption from wastewater (Brinza et al., 2007;
Ummalyma et al., 2018). Biosorption studies of Scenedesmus sp. showed the potential of said
species to remove heavy metals namely Cu (73-98%), Zn (65-98%), Cr (81-96%) and Pb (75-
98%) from tannery wastewater under laboratory conditions (Ajayan et al., 2015). Similarly, the
biomass of Chara aculeolate removed Cd, Pb, and Zn via biosorption at the rate of 23 mgg-1,
105.3 mgg-1 and 15.2 mgg-1 respectively from municipal wastewater (Sooksawat et al., 2016).
Moreover, S. armatus and C. vulgaris showed bioaccumulation efficiency for Cd2+ and Pb2+ by
89.96% and 88.98%, respectively (Zabochnicka-Świątek and Rygała, 2017). Similarly,
Cladophora glomerate and Oedogonium westii showed the removal of Cd and Ni by 80% and
66.3%, respectively from industrial wastewater (Khan et al., 2017). Interesting results were
obtained for self-flocculating microalga C. vulgaris JSC-7 which was able to remove Cd and Zn
(60-80% respectively) more efficiently as compared to the non-flocculating similar strain.
Moreover, an increase in the photosynthetic pigments and growth was observed under heavy
metal stress, showing its excellent heavy metal tolerance ability (Alam et al., 2015).

Heavy metal accumulation is associated with toxicity and ROS production where Pb, Cu,
Cd, and Ti greatly reduce the growth and pigment content of microalgae. However, in some
cases, an increase in the protein and lipid content was also observed. Heavy metal-containing
wastewater poses many adverse effects and is not suitable for human consumption.
Phycoremediation of heavy metal-containing wastewater is an eco-friendly and cost-effective
approach owing to the remarkable potential of microalgae to remove high levels of heavy metals
from wastewater.

2.2. Impact of pH
Invasion of pathogenic microbes (predators or competitors) in the open pond cultivation
of microalgae causes contamination of microalgae and poses a major challenge during
cultivation. There are several factors that can influence the load of invading organisms. For
instance, microalgae may produce some extracellular compounds to inhibit the competing
organisms resultantly dominating the environment. Some secretory metabolites can even help the
microalgae to modify the pH of the media (Shahid et al., 2019) which helps to outcompete the
invading organisms. Hence, pH is one of the most important parameters which can be employed
to inhibit or outcompete the invading organisms. Alteration in pH has been reported to improve

12
culture densities by decreasing the microalgal contaminants (Bartley et al., 2014). Additionally,
pH plays a vital role in algal cultivation as it is responsible for nutrient and CO2 availability and
solubility. Moreover, it is known to affect the activity of various enzymes to enhance the
triglyceride accumulation (Juneja et al., 2013; Ying et al., 2014). Optimum pH for microalgae
cultivation usually ranges from 7.0-7.6 (neutral pH) but, pH tolerance is species-specific and
maximum biomass productivities for S. obliquus and Ettlia sp. were observed at pHs 7.0 and 8.5,
respectively (Chen et al., 2017). The pH of the growth media influences the microbial enzyme
activity and solubility of environmental micro-pollutants. Therefore, pH variation determines the
fate of pollutants during bioreactor treatments. Furthermore, acidic to neutral pH values are
significant for improved pharmaceutical degradation (Grandclement et al., 2017).

Cultivation of C. vulgaris at pH ranging from 3-11 represented its ability to act

differently under specific pH as highest biomass productivity (0.541 days-1) and lipid content of
53% was observed at pH 7.5 while, pH 9.5 found to be suitable for cell aggregation (Sakarika
and Kornaros, 2016). Interestingly, in the case of N. oleoabundans, pH 9.5 supported maximum
cell growth (1.04 gL-1), lipid content (151.2 mgg-1) and lipid productivity (19.1 mgL-1day-1)
under oxygen stress (Peng et al., 2017). S. abundans was able to tolerate a large range of pH (5-
8) however, pH 8 was optimum to obtain the highest growth rate of 769 mgL-1 and pH 6 favored
lipid concentration to 179 mgL-1 (Mandotra et al., 2016). Similarly, pH alteration enhanced the
self-flocculation ability of Dunaliella sp. when 2 or 6 N NaOH was added in the culture media
(Byrd and Burkholder, 2017). These studies indicated that pH manipulation can be employed to
enhance the algal metabolite content and biomass productivity by outcompeting the pathogens.
However, more detailed studies are required to elucidate the impact of wastewater-derived pH on
biomass productivity and metabolite content.

3. Binary culture to enhance microalgal phycoremediation of wastewater

It is difficult to maintain pure cultures under field conditions, which is even more
difficult when using ponds receiving wastewater. Hence, recently researchers focused on the
cultivation of mixed cultures. Binary cultures in the form of consortia (microalgae-microalgae or
microalgae-bacteria) have been reported extensively for enhanced wastewater treatment due to
their higher nutrient removal ability with enhanced biomass production (Table 3). Polycultures
allow us to develop robust biological systems for wastewater treatment as they can be combined
with various metabolic processes to enable themselves to survive under environmental stress
conditions. Moreover, integrated consortia can uptake nutrients at a higher rate (Johnson and
Admassu, 2013; Rawat et al., 2011; Renuka et al., 2013) because one strain can remove nitrogen
and another can remove heavy metals specifically. These consortia have several advantages
including (i) contamination and predator resistances due to production of allelochemicals (ii)
enhanced nutrient consumption; ensuring sufficient nutrient supply during whole process, (iii)
development of settleable system for flocculation thus, eliminating limitations of harvesting, and
(iv) enhanced viability of phycoremediation; as loss of one microbe is compensated by other
species. However, it faces some constraints as it’s difficult to develop robust consortia as a
variety of combinations are possible. Moreover, maintenance of consortia for longer periods
especially in an open-pond system proven to be challenging (Gonçalves et al., 2017).

3.1. Microalgae-bacteria (MB) consortia

The symbiotic relationship of microalgae and bacteria may be in the form of
commensalism, mutualism or parasitism. In general, bacteria heterotrophically produce CO2 and
other important nutrients that are consumed by microalgae for their growth (Zhu et al., 2019). In
return, oxygen produced by microalgae during photosynthesis is valuable for bacteria (Rashid et
al., 2018). Moreover, bacteria provide growth-promoting hormones and vitamin B to microalgae
which are necessary for growth (Fuentes et al., 2016). Additionally, this symbiotic relationship
protects the microalgae from other invading species. On the other hand, bacteria can damage the
microalgal cell wall to utilize intracellular nutrient (Magdouli et al., 2016). This property is of
special interest during harvesting stages where cell rupturing is required to obtain the desired
product thus, reducing cost and time of downstream processing in the biorefinery. The MB-
consortia also consume the dead algal cells as a nutrient source (Ramanan et al., 2016). Excretion
of special chemicals by bacteria and microalgae may also suppress the growth of each other
(López-Serna et al., 2019). This property can be useful to reduce sterilization cost as it eliminates
the contamination chances (Kouzuma and Watanabe, 2015).

The MB-consortia have shown to be effective for removal of phosphorus (35-88%),

nitrogen (43-89%) and carbon (59-80%) respectively from municipal wastewater in a lab-scale
photobioreactor (Lee et al., 2015). Moreover, a marine algal-microbial (Picochlorum sp,
Pseudomonas sp., and Chitrinomycetes) consortium was developed for the treatment of marine

14
aquaculture effluents which showed up to 95% of removal efficiency in 4-5 hours (Babatsouli et
al., 2015). Improvement in total phosphorus and total nitrogen removal by 46% and 12% from
domestic wastewater was observed in batch reactors cultivated with the MB-consortia (Tang et
al., 2016). The MB-consortium based on B. licheniformis + C. vulgaris showed excellent
efficiency to remove COD (86.5%), TN (88.9%) and TP (80.2%) from synthetic wastewater (Ji
et al., 2018). Similarly, the symbiotic relationship of Chlorella and Proteobacteria removed 72%
TN in an outdoor cultivation system while 100% TP and 83% Zn removal efficiency were
observed in an indoor cultivation system using piggery wastewater (García et al., 2017).

3.2. Microalgae-microalgae consortia

In order to minimize the issues and challenges regarding the microalgal cultivation, one
promising alternative is to cultivate multiple algal species which have a synergistic impact on
one another to enhance the production and productivities as diverse communities may enhance
biomass specific lipid production when compared to corresponding monocultures (Stockenreiter
et al., 2016). Additionally, diversity correlates with the biomass stability which is an important
requirement of mass cultivation (Nalley et al., 2014). Moreover, a prodigious variety of traits by
primary producer communities relates to higher diversity (Stockenreiter et al., 2016).

More than 96% of nutrient removal efficiency along-with 6.82% lipid content and 9.2-
17.8 tonnes year-1ha-1 of biomass productivity was achieved using polyculture of 15 native-algal
strains (Chinnasamy et al., 2010). Mixed microalgal cultures predominantly consisting of
Chlorella with small amounts of Scenedesmus could remove 31 mgL-1 of phosphorus and 481
mgL-1 of nitrogen from textile wastewater when cultivated under photoautotrophic conditions
(Huy et al., 2018). Polyculture of Chlorella sp. displayed the ability to removal 100% nitrates
from swine wastewater and accumulated high lipid and protein content in the ratio of 59% and
34% respectively. However, lipid content only accounts for 3% of total biomass content
(Michelon et al., 2016).

3.3. Myco-algal consortia

Naturally, microalgae live in association with fungi in the form of lichens. This strategy
benefits both partners; fungi obtain essential nutrients and sugars from algae and in return
provide protection to microalgae from abiotic stress. It is proposed that this association can act as
a self-sufficient organization to improve the overall performance and economics of the integrated
microalgal industry at large scale (Ummalyma et al., 2017). Additionally, fungal pellets may act
as natural coagulants and ultimately helping the microalgal flocculation, hence making
harvesting easier. These pellets also have the potential of treating wastewater by entrapping
sludge solids (Ummalyma et al., 2017). Furthermore, 30% of the total fungal biomass consists of
lipids, making them suitable biodiesel producer candidates (Zhou et al., 2013).

The mixed culture of C. vulgaris and Mucor indicus reduced the phosphate and total
ammonia and nitrogen (TAN) to almost undetectable levels from synthetic aquaculture
wastewater. This process also enabled the flocculation of 860 mg DW of myco-algal biomass
(Barnharst et al., 2018). Biogas up-gradation and domestic sewage wastewater treatment were
simultaneously performed by algal-fungal co-cultivation which removed TP and COD by 81%
(Xu et al., 2017). Similarly, co-cultivation of C. vulgaris with Ganoderma lucidum reduced TP,
TN and COD concentrations by 84%, 74%, and 79% respectively from swine wastewater
treatment. Moreover, an association of P. subcapitata and G. lucidum successfully removed CO2
with 84% efficiency for biogas up-gradation (Guo et al., 2017). These studies have clearly
demonstrated that myco-algal associations have the potential to enhance the wastewater
treatment potential and fungal pellets can contribute towards easier harvesting of the algal
biomass. However, detailed studies are required to evaluate the impact of myco-algal
associations on the metabolite content of the microalgae. Because the presence of fungi in the
culture media can substantially modify the extracellular and intracellular algal metabolites which
may improve or even lower the content of any desired molecule. Moreover, the fungal pellets
may also interfere with the post-harvesting processing of the algal biomass for the extraction of
any desired organic compounds.

4. Carbon dioxide fixation

According to 2014 statistics, the total emission of CO2 was 6,870 MMT (Million metric
tons), equivalent to 81% of the world’s GHG emission (Wilbanks and Fernandez, 2014) and
major global warming contributor. Industrial processes contributed to 21% of GHG emissions
(Cheah et al., 2015). Microalgae are believed to have the potential to fix the atmospheric carbon
at the highest rate when compared with any other photosynthetic system (Gill et al., 2016; Gill et
al., 2013). Algae have been extensively studied for GHG mitigation by reducing the CO2 content

16
in biosphere through photosynthetic fixation (Abid et al., 2017). Their CO2 sequestration
efficiency is 10-50 times higher than terrestrial plants (Abid et al., 2017; Cheng et al., 2013).
Microalgae can convert CO2 from industrial gas and atmosphere into high chemical energy-
containing organic biomass like carotenoid, bioethanol, acetone and lipids (Zhu et al., 2017).
Different microalgae namely S. obliquus, C. pyrenoidosa, C. reinhardtii, Chlorococcum littorale,
and D. terteolacta have shown the ability of HCO3-1 and CO2 utilization because they harbor
external carbonic anhydrase (Zhou et al., 2017). Immobilized Nannochloropsis sp. mitigated >99
% CO2 in secondary POME (Palm Oil Mill Effluent) with concomitant lipid production of 0.35
gL-1 lipid production which was 1.41-fold higher when compared to the control (Cheirsilp et al.,
2017). Cultivation of S. obliquus in beer wastewater supplemented with 5% CO2 promoted cell
growth and lipid accumulation (Wu et al., 2017b). Microalgae-based sludge treatment reduced
CO2 emission by 22-54% and 43-103% during the whole year and in summer, respectively
(Nordlander et al., 2017). Phycoremediation of palm oil mill secondary effluents by immobilized
Nannochloropsis sp. showed >99% CO2 mitigation efficiency (Cheirsilp et al., 2017).

The CO2 content may manipulate the algal metabolite content. For instance, higher fatty
acid content was achieved in C. reinhardtii, S. obliquus, C. minutissima, and D. tertiolecta by
exposing them to enhanced CO2 concentrations (Zhu et al., 2017). In another study, involving
eight locally isolated microalgal strains, cultivation was conducted under 20% CO2 feeding
(Hussain et al., 2017), and maximum biomass production of 1.4 gL-1 for strain UMN268 was
observed and it was correlated with nutrient uptake. Exposure of Asterarcys quadricellulare
and C. sarokiniana to high levels of CO2 resulted in an increase in carbohydrate content of total
dry cell mass by 55-71% (Varshney et al., 2018). Similarly, S. bajacalifornicus supplemented
with 25% CO2 accumulated lipid and carbohydrate by 25.81% and 26.19%, respectively (Patil
and Kaliwal, 2017). These studies have clearly indicated that CO2 concentration can manipulate
the content and the composition of the algal metabolites and this manipulation may be employed
to achieve the desired metabolite content in selected microalgal strains. However, the impact of
CO2 content derived from wastewater (the soluble form of CO2) needs to be carefully evaluated
in future through wastewater-oriented studies because wastewater from different sources may
have different levels of soluble CO2 depending upon nature, geographic position and origin of
the wastewater. Figure 3 shows a schematic diagram of microalgal cultivation and processing as
a carbon neutral process.
5 Wastewater integrated algal-biorefinery for bioproducts

Circular bio-economy is an emerging concept, focusing on the sustainable production,

conversion, and utilization of renewable resources into value-added products. Photosynthetic
organisms such as microalgae are the focal point in developing closed-loop systems due to their
eco-friendly and versatile properties. Wastewater integrated closed-biorefinery contributes to
process economy and sustainability by resource recovery and by reducing the ecological
footprint (Javed et al., 2019; Mohan et al., 2019a). Wastewater cultivated algae are rich source of
primary (carbohydrates, proteins, lipids) and secondary (pigments, anti-oxidants) metabolites
that could be exploited to produce biofuels, bio-polymers, biofertilizers, nutraceuticals,
food/health grade compounds, enzymes, feed supplements etc. and the treated wastewater can be
reutilized for the agricultural or industrial purposes (Mohan et al., 2019a).

5.1. Algae as fuel source

Algae are known to accumulate 30-80 % of lipids which normally consist of 90-95 % of
triacylglycerides. Wastewater cultivation is a cost-effective method to enhance biomass
production and to modify lipid content and fatty acid (FA) composition. Most of the algal strains
such as Chlorella produce FA ranging from C16-C18 which are suitable for biodiesel production;
similar in properties to traditional fossil-based diesel. Ethanol is another important fuel and a
chemical source. It can be produced from syngas or directly by fermenting high carbohydrate
(>40%) containing algal strains. Algal species like Chlamydomonas, Spirulina, Euglena,
Chlorella, Scenedesmus, and Dunaliella have been extensively studied for bioethanol
production. Various studies indicated the potential of wastewater grown algae to accumulate
carbohydrates (Mehar et al., 2019). Residual biomass (lipid-free, carbohydrate rich) can also be
utilized for bioethanol fermentation (Shahid et al., 2019) and doesn’t require laborious
pretreatments as the case with plant biomass. Carbohydrate content of algae can also be used to
produce bio-butanol, bio-methane, biogas, and syngas.

5.2. Algae as high-value nutraceutical/health supplement

Some algal strains tend to produce health related FA like omega-3, omega-6 and
docosahexaenoic acid (DHA) as major algal-based FA. They are non-toxic and more stable as
18
compared to fish DHA (Kumar and Singh, 2019). Cyanobacteria and algae are the natural
sources for carotenoids and diverse carotenoids including astaxanthin and lutein. Production of
these compounds can be enhanced through abiotic factor manipulation. Wastewater grown algae
can be utilized for this purpose and provides a cost-effective sustainable alternative. Astaxanthin
has vast applications in medical sciences due to its anti-oxidant, anti-inflammatory, anti-cancer,
and anti-aging properties. Moreover, it also improves the nervous system, respiratory system,
fertility, and digestive system. Wastewater integrated algal-growth has been suggested for
improved astaxanthin production. This impact has been reviewed in detail by Shah et al., (Shah,
2019). Similarly, other carotenoids have shown various nutraceutical properties. Cyanobacteria
(group of algae) produce large amounts of industrially and nutraceutically important compounds
called phycobilins (Pancha et al., 2019) which are colored protein-pigment complexes having
antimicrobial, anticancer, and antioxidant properties. They can also be used to produce protein-
rich energy drinks and can be applied has food colors.

Wastewater-cultivated algae could be used as animal feed, biofertilizer, and cosmetic

agents. Biochar produced by algal pyrolysis is rich source of nutrients and can be utilized as
biofertilizer in agriculture. Spirulina, Chlorella, Euglena, Tetraselmis, Synechococcus,
Nannochloropsis have been utilized for human consumption as well as aquaculture and animal
feed supplement due to their high protein content and nutritional value (Shahid et al., 2020).
Algal extracts have been widely used in cosmetic industry as skin and hair protectants due to
their anti-oxidant, anti-irritant, anti-aging, sun-protecting and tissue regenerating abilities.
Chondrus, Chlorella, Dunaliella, Nannochloropsis, Spirulina have been widely utilized for
commercial cosmetic products (Javed et al., 2019; Shahid et al., 2020). Figure 4 shows
production and processing of algal biomass to produce diverse products in an algal biorefinery.

6 Conclusion, prospects and recommendations

Microalgae are a remarkable biological resource to produce biofuels and value-added

products at industrial scale in an eco-friendly manner. However, the cost associated with algal
biomass production and processing questions the commercial viability of the process.
Fortunately, microalgal cultivation in wastewater offers a promising alternative to deal with the
higher operational cost. Because wastewater does not only provide a low-cost growth media for
algal cultivation but also provides cost-benefits associated with the wastewater treatment.
Moreover, the unbalanced ratios of nutrients on the microalgae may exert abiotic stresses on
algal biomass diverting the metabolic pathways to produce either more protein or more lipids and
pigments which may add additional value. In this regard, a biorefinery approach is feasible
where a spectrum of marketable products may be obtained from the single process. Wastewater
is a major global problem especially in heavily populated areas and must be recycled or treated
before disposal. On the other hand, wastewater is a rich source of nutrients and consumption of
these nutrients by microalgae (phycoremediation) not only enhance productivities of biomass but
other value-added products (lipids, pigments, proteins, carbohydrates, etc.). Moreover, the
nutrient removal efficiency of microalgae can be enhanced by applying binary cultures that may
consist of microalgae-microalgae, microalgae-bacteria or microalgae-fungi associations. These
associations also improve the flocculation ability while avoiding the contamination chances and
may improvise the natural metabolite content of microalgae. However, detailed studies are
required in the future to study the impact of wastewater-derived stress on the biomass
productivity of consortia and the post-harvesting processing of the biomass along with the
metabolite content. Moreover, stress-responsive genes should be identified in the future which
would be later targeted to engineer the selected microalgal strains to achieving industrial
robustness in algal biorefineries. Keeping in view the progress made in algal research, recent
scenario and prospects, the following recommendations are made to achieve commercial
robustness.
 Cultivation: the microalgal cultivation systems need to be improved by developing low-
cost growth media or using wastewaters as growth media to reduce the cost.
Bioprospecting is required to isolate indigenously successfully adapted strains for
regional or territorial applications. The strains adapted to grow at odd pH values (low or
high) may be selected to outcompete the contaminating microbes at large-scale
cultivation.
 Harvesting: harvesting involves 20-30% cost in the algal biomass production and
processing which can be lowered by developing robust harvesting techniques. Bio-
flocculation and self-flocculation seem attractive harvesting approaches. Hence, isolation
of self-flocculating strains and robust bio-flocculation would be required to develop in
the future.

20
  Strain development: unlike Saccharomyces cerevisiae and Escherichia coli, the metabolic
engineering systems are not fully established to develop engineered robust algal strains.
Hence, it is recommended that multi-OMICs, synthetic and system biology-based
research should be focused to elucidate and engineer the metabolic pathways of the
selected algal strains.
 Downstream processing: the processing of microalgal biomass into products is tedious
and expensive. It would be better if a multi-product approach is adopted for the algal
biorefinery. It would be required to conduct detailed studies to elucidate the technical and
economical impact of using residual algal biomass for traditional applications in energy,
environment and agriculture sectors after extracting the high-value products.
 Cost-energy-environment input-output balance: more detailed studies are required to
study the balance ratios of the cost-energy-environment triangle especially when it comes
to a large-scale multi-product algal biorefinery.

Acknowledgments
Authors are highly obliged to 1000 Talent Plan of Sichuan (China) government and Higher
Education Commission Pakistan for their financial support.
Conflict of interest
Authors declare that they have no competing interests.

References

Abid A, Saidane F, Hamdi M. Feasibility of carbon dioxide sequestration by Spongiochloris sp

microalgae during petroleum wastewater treatment in airlift bioreactor. Bioresour.
Technol. 2017; 234: 297-302.
Affam AC, Chaudhuri M, Kutty SRM. Comparison of five advanced oxidation processes for
degradation of pesticide in aqueous solution. Bullet. Chem. React. Eng. Catalys. 2018;
13: 179-186.
Afshan S, Ali S, Bharwana SA, Rizwan M, Farid M, Abbas F, et al. Citric acid enhances the
phytoextraction of chromium, plant growth, and photosynthesis by alleviating the
 oxidative damages in Brassica napus L. Environ. Sci. Pollut. Res. 2015; 22: 11679-
11689.
Afzal I, Shahid A, Ibrahim M, Liu T, Nawaz M, Mehmood MA. Microalgae: A promising
feedstock for energy and high-value products. Algae Based Polymers, Blends, and
Composites, 2017, pp. 55.
Ahmad T, Aadil RM, Ahmed H, ur Rahman U, Soares BC, Souza SL, et al. Treatment and
utilization of dairy industrial waste: A review. Trends Food Sci. Technol. 2019.
Ajayan KV, Selvaraju M, Unnikannan P, Sruthi P. Phycoremediation of tannery wastewater
using microalgae Scenedesmus species. Int. J. Phytoremediation 2015; 17: 907-916.
Alam MA, Wan C, Zhao X-Q, Chen L-J, Chang J-S, Bai F-W. Enhanced removal of Zn2+ or
Cd2+ by the flocculating Chlorella vulgaris JSC-7. J. Hazard. Mater. 2015; 289: 38-45.
Ali I, Peng C, Khan ZM, Naz I, Sultan M, Ali M, et al. Overview of microbes based fabricated
biogenic nanoparticles for water and wastewater treatment. J. Environ. Manage. 2019;
230: 128-150.
Álvarez-Díaz P, Ruiz J, Arbib Z, Barragán J, Garrido-Pérez M, Perales J. Freshwater microalgae
selection for simultaneous wastewater nutrient removal and lipid production. Algal Res.
2017.
Anand J, Arumugam M. Enhanced lipid accumulation and biomass yield of Scenedesmus
quadricauda under nitrogen starved condition. Bioresour. Technol. 2015; 188: 190-194.
Antal TK, Matorin DN, Kukarskikh GP, Lambreva MD, Tyystjärvi E, Krendeleva TE, et al.
Pathways of hydrogen photoproduction by immobilized Chlamydomonas reinhardtii cells
deprived of sulfur. Int. J. Hydrog. Energy 2014; 39: 18194-18203.
Apandi N, Mohamed RMSR, Al-Gheethi A, Gani P, Ibrahim A, Kassim AHM. Scenedesmus
biomass productivity and nutrient removal from wet market wastewater, a bio-kinetic
study. Waste Biomass Valorization 2019; 10: 2783-2800.
Aziz A, Basheer F, Sengar A, Khan SU, Farooqi IH. Biological wastewater treatment (anaerobic-
aerobic) technologies for safe discharge of treated slaughterhouse and meat processing
wastewater. Sci. Total Environ. 2019.
Babatsouli P, Fodelianakis S, Paranychianakis N, Venieri D, Dialynas M, Kalogerakis N. Single
stage treatment of saline wastewater with marine bacterial–microalgae consortia in a
fixed-bed photobioreactor. J. Hazard. Mater. 2015; 292: 155-163.

22
Barnharst T, Rajendran A, Hu B. Bioremediation of synthetic intensive aquaculture wastewater
by a novel feed-grade composite biofilm. Int. Biodeterior. Biodegrad. 2018; 126: 131-
142.
Bartley ML, Boeing WJ, Dungan BN, Holguin FO, Schaub T. pH effects on growth and lipid
accumulation of the biofuel microalgae Nannochloropsis salina and invading organisms.
J. Appl. Phycol. 2014; 26: 1431-1437.
Batool S, Hussain A, Iqbal MA, Javid A, Ali W, Bukhari SM, et al. Implication of highly metal-
resistant microalgal-bacterial co-cultures for the treatment of simulated metal-loaded
wastewaters. Int. Microbiol. 2019; 22: 41-48.
Benítez MB, Champagne P, Ramos A, Torres AF, Ochoa-Herrera V. Wastewater treatment for
nutrient removal with Ecuadorian native microalgae. Environ. Technol. 2019; 40: 2977-
2985.
Bhattacharya S, Pramanik SK, Gehlot PS, Patel H, Gajaria T, Mishra S, et al. Process for
preparing value-added products from microalgae using textile effluent through a
biorefinery approach. ACS Sustain. Chem. Eng. 2017.
Bilal M, Shah JA, Ashfaq T, Gardazi SMH, Tahir AA, Pervez A, et al. Waste biomass
adsorbents for copper removal from industrial wastewater—a review. J. Hazard. Mater.
2013; 263: 322-333.
Brinza L, Dring MJ, Gavrilescu M. Marine micro and macro algal species as biosorbents for
heavy metals. Environ. Eng. Manag. J. 2007; 6.
Butkovskyi A, Leal LH, Zeeman G, Rijnaarts H. Micropollutants in source separated wastewater
streams and recovered resources of source separated sanitation. Environ. Res. 2017; 156:
434-442.
Byrd SM, Burkholder JM. Environmental stressors and lipid production in Dunaliella spp. II.
nutrients, pH, and light under optimal or low salinity. J. Exp. Mar. Biol. Ecol. 2017; 487:
33-44.
Cakmak T, Angun P, Demiray YE, Ozkan AD, Elibol Z, Tekinay T. Differential effects of
nitrogen and sulfur deprivation on growth and biodiesel feedstock production of
Chlamydomonas reinhardtii. Biotechnol. Bioeng. 2012; 109: 1947-1957.
Calicioglu O, Demirer GN. Carbon-to-Nitrogen and Substrate-to-Inoculum ratio adjustments can
improve co-digestion performance of microalgal biomass obtained from domestic
wastewater treatment. Environ. Technol. 2017: 1-23.
Cheah WY, Show PL, Chang J-S, Ling TC, Juan JC. Biosequestration of atmospheric CO2 and
flue gas-containing CO2 by microalgae. Bioresour. Technol. 2015; 184: 190-201.
Cheirsilp B, Thawechai T, Prasertsan P. Immobilized oleaginous microalgae for production of
lipid and phytoremediation of secondary effluent from palm oil mill in fluidized bed
photobioreactor. Bioresour. Technol. 2017; 241: 787-794.
Chen B, Wan C, Mehmood MA, Chang J-S, Bai F, Zhao X. Manipulating environmental stresses
and stress tolerance of microalgae for enhanced production of lipids and value-added
products-A review. Bioresour. Technol. 2017.
Chen G, Zhao L, Qi Y. Enhancing the productivity of microalgae cultivated in wastewater
toward biofuel production: A critical review. Appl. Energ. 2015; 137: 282-291.
Chen X, Hu Z, Qi Y, Song C, Chen G. The interactions of algae-activated sludge symbiotic
system and its effects on wastewater treatment and lipid accumulation. Bioresour.
Technol. 2019; 292: 122017.
Cheng J, Huang Y, Feng J, Sun J, Zhou J, Cen K. Improving CO2 fixation efficiency by
optimizing Chlorella PY-ZU1 culture conditions in sequential bioreactors. Bioresour.
Technol. 2013; 144: 321-327.
Cheng J, Qiu H, Chang Z, Jiang Z, Yin W. The effect of cadmium on the growth and antioxidant
response for freshwater algae Chlorella vulgaris. SpringerPlus 2016; 5: 1290.
Chinnasamy S, Bhatnagar A, Hunt RW, Das K. Microalgae cultivation in a wastewater
dominated by carpet mill effluents for biofuel applications. Bioresour. Technol. 2010;
101: 3097-3105.
Chisti Y. Biodiesel from microalgae. Biotechnol. Adv. 2007; 25: 294-306.
Chokshi K, Pancha I, Ghosh A, Mishra S. Nitrogen starvation-induced cellular crosstalk of ROS-
scavenging antioxidants and phytohormone enhanced the biofuel potential of green
microalga Acutodesmus dimorphus. Biotechnol. Biofuel 2017; 10: 60.
Choudhary P, Prajapati SK, Malik A. Screening native microalgal consortia for biomass
production and nutrient removal from rural wastewaters for bioenergy applications. Ecol.
Eng. 2016; 91: 221-230.

24
Crini G, Lichtfouse E. Advantages and disadvantages of techniques used for wastewater
treatment. Environ. Chem. Letters 2019; 17: 145-155.
Crini G, Lichtfouse E, Wilson LD, Morin-Crini N. Conventional and non-conventional
adsorbents for wastewater treatment. Environmental Chemistry Letters 2019; 17: 195-
213.
Daneshvar E, Antikainen L, Koutra E, Kornaros M, Bhatnagar A. Investigation on the feasibility
of Chlorella vulgaris cultivation in a mixture of pulp and aquaculture effluents:
Treatment of wastewater and lipid extraction. Bioresour. Technol. 2018; 255: 104-110.
Daneshvar E, Zarrinmehr MJ, Koutra E, Kornaros M, Farhadian O, Bhatnagar A. Sequential
cultivation of microalgae in raw and recycled dairy wastewater: microalgal growth,
wastewater treatment and biochemical composition. Bioresour. Technol. 2019; 273: 556-
564.
Das A, Osborne JW. Bioremediation of heavy metals. Nanotechnology, Food Security and Water
Treatment. Springer, 2018, pp. 277-311.
Dasan YK, Lam MK, Yusup S, Lim JW, Lee KT. Life cycle evaluation of microalgae biofuels
production: Effect of cultivation system on energy, carbon emission and cost balance
analysis. Sci. Total Environ. 2019; 688: 112-128.
Daud M, Nafees M, Ali S, Rizwan M, Bajwa RA, Shakoor MB, et al. Drinking water quality
status and contamination in Pakistan. BioMed Res. Int. 2017; 2017.
de Alva MS, Pabello VML, Ledesma MTO, Gómez MJC. Carbon, nitrogen, and phosphorus
removal, and lipid production by three saline microalgae grown in synthetic wastewater
irradiated with different photon fluxes. Algal Res. 2018; 34: 97-103.
De Francisci D, Su Y, Iital A, Angelidaki I. Evaluation of microalgae production coupled with
wastewater treatment. Environ. Technol. 2018; 39: 581-592.
Del Río E, García-Gómez E, Moreno J, Guerrero MG, García-González M. Microalgae for oil.
assessment of fatty acid productivity in continuous culture by two high-yield strains,
Chlorococcum oleofaciens and Pseudokirchneriella subcapitata. Algal Res. 2017; 23:
37-42.
Deng X-Y, Gao K, Zhang R-C, Addy M, Lu Q, Ren H-Y, et al. Growing Chlorella vulgaris on
thermophilic anaerobic digestion swine manure for nutrient removal and biomass
production. Bioresour. Technol. 2017; 243: 417-425.
Diniz GS, Silva AF, Araújo OQ, Chaloub RM. The potential of microalgal biomass production
for biotechnological purposes using wastewater resources. J. Appl. Phycol. 2017; 29:
821-832.
Dvořák L, Lederer T, Jirků V, Masák J, Novák L. Removal of aniline, cyanides and
diphenylguanidine from industrial wastewater using a full-scale moving bed biofilm
reactor. Process Biochem. 2014; 49: 102-109.
Ejraei A, Aroon MA, Saravani AZ. Wastewater treatment using a hybrid system combining
adsorption, photocatalytic degradation and membrane filtration processes. J. Water
Process. Eng. 2019; 28: 45-53.
Enaime G, Baçaoui A, Yaacoubi A, Wichern M, Lübken M. Olive mill wastewater pretreatment
by combination of filtration on olive stone filters and coagulation–flocculation. Environ.
Technol. 2019; 40: 2135-2146.
Eyvaz M, Arslan S, Gürbulak E, Yüksel E. Textile materials in liquid filtration practices: Current
status and perspectives in water and wastewater treatment. Textiles for Advanced
Applications. InTech, 2017.
Fan J, Chen Y, Zhang TC, Ji B, Cao L. Performance of Chlorella sorokiniana-activated sludge
consortium treating wastewater under light-limited heterotrophic condition. Chem. Eng.
J. 2020; 382: 122799.
Ferreira A, Ribeiro B, Ferreira AF, Tavares ML, Vladic J, Vidović S, et al. Scenedesmus
obliquus microalga‐based biorefinery–from brewery effluent to bioactive compounds,
biofuels and biofertilizers–aiming at a circular bioeconomy. Biofuels, Bioproducts and
Biorefining 2019; 13: 1169-1186.
Ferreira A, Ribeiro B, Marques PA, Ferreira AF, Dias AP, Pinheiro HM, et al. Scenedesmus
obliquus mediated brewery wastewater remediation and CO2 biofixation for green energy
purposes. J. Clean. Prod. 2017; 165: 1316-1327.
Fuentes JL, Garbayo I, Cuaresma M, Montero Z, González-del-Valle M, Vílchez C. Impact of
microalgae-bacteria interactions on the production of algal biomass and associated
compounds. Mar. Drugs 2016; 14: 100.
Ganeshkumar V, Subashchandrabose SR, Dharmarajan R, Venkateswarlu K, Naidu R, Megharaj
M. Use of mixed wastewaters from piggery and winery for nutrient removal and lipid
production by Chlorella sp. MM3. Bioresour. Technol. 2018; 256: 254-258.

26
Gani P, Sunar NM, Matias-Peralta H, Mohamed RMSR, Latiff AAA, Parjo UK. Extraction of
hydrocarbons from freshwater green microalgae (Botryococcus sp.) biomass after
phycoremediation of domestic wastewater. Int. J. Phytoremediation 2017; 19: 679-685.
Gao F, Li C, Yang ZH, Zeng GM, Mu J, Liu M, et al. Removal of nutrients, organic matter, and
metal from domestic secondary effluent through microalgae cultivation in a membrane
photobioreactor. J. Chem. Technol. Biotechnol. 2016; 91: 2713-2719.
Gao F, Peng Y-Y, Li C, Yang G-J, Deng Y-B, Xue B, et al. Simultaneous nutrient removal and
biomass/lipid production by Chlorella sp. in seafood processing wastewater. Sci. Total
Environ. 2018; 640: 943-953.
García D, Posadas E, Grajeda C, Blanco S, Martínez-Páramo S, Acién G, et al. Comparative
evaluation of piggery wastewater treatment in algal-bacterial photobioreactors under
indoor and outdoor conditions. Bioresour. Technol. 2017; 245: 483-490.
Gautam PK, Singh A, Misra K, Sahoo AK, Samanta SK. Synthesis and applications of biogenic
nanomaterials in drinking and wastewater treatment. J. Environ. Manage. 2019; 231: 734-
748.
Gill SS, Mehmood MA, Ahmad N, Ibrahim M, Rashid U, Ali S, et al. Strain selection, growth
productivity and biomass characterization of novel microalgae isolated from fresh and
wastewaters of upper Punjab, Pakistan. Front. Life Sci. 2016; 9: 190-200.
Gill SS, Mehmood MA, Rashid U, Ibrahim M, Saqib A, Tabassum MR. Waste-water treatment
coupled with biodiesel production using microalgae: a bio-refinery approach. Pakistan J.
Life Soc. Sci. 2013; 11: 179-189.
Giordano M, Raven JA. Nitrogen and sulfur assimilation in plants and algae. Aquat. Bot. 2014;
118: 45-61.
Godfray HCJ, Beddington JR, Crute IR, Haddad L, Lawrence D, Muir JF, et al. Food security:
the challenge of feeding 9 billion people. Science 2010; 327: 812-818.
Gonçalves AL, Pires JC, Simões M. A review on the use of microalgal consortia for wastewater
treatment. Algal Res. 2017; 24: 403-415.
Goswami G, Makut BB, Das D. Sustainable production of bio-crude oil via hydrothermal
liquefaction of symbiotically grown biomass of microalgae-bacteria coupled with
effective wastewater treatment. Sci. Rep. 2019; 9: 1-12.
Grandclement C, Seyssiecq I, Piram A, Wong-Wah-Chung P, Vanot G, Tiliacos N, et al. From
the conventional biological wastewater treatment to hybrid processes, the evaluation of
organic micropollutant removal: a review. Water Res. 2017.
Guldhe A, Ansari FA, Singh P, Bux F. Heterotrophic cultivation of microalgae using aquaculture
wastewater: A biorefinery concept for biomass production and nutrient remediation. Ecol.
Eng. 2017; 99: 47-53.
Guo G, Cao W, Sun S, Zhao Y, Hu C. Nutrient removal and biogas upgrading by integrating
fungal–microalgal cultivation with anaerobically digested swine wastewater treatment. J.
Appl. Phycol. 2017; 29: 2857-2866.
Guo R, Cai X, Liu H, Yang Z, Meng Y, Chen F, et al. In situ growth of metal–organic
frameworks in three-dimensional aligned lumen arrays of wood for rapid and highly
efficient organic pollutant removal. Environ. Sci. Technol. 2019; 53: 2705-2712.
Gupta SK, Ansari FA, Shriwastav A, Sahoo NK, Rawat I, Bux F. Dual role of Chlorella
sorokiniana and Scenedesmus obliquus for comprehensive wastewater treatment and
biomass production for bio-fuels. J. Clean. Prod. 2016; 115: 255-264.
Hariz HB, Takriff MS. Palm oil mill effluent treatment and CO2 sequestration by using
microalgae—sustainable strategies for environmental protection. Environ. Sci. Pollut.
Res. 2017: 1-32.
Hazman NAS, Yasin NHM, Takriff MS, Hasan HA. Integrated palm oil mill effluent treatment
and CO2 sequestration by microalgae. Sains Malaysiana 2018; 47: 1455-1464.
Hena S, Fatimah S, Tabassum S. Cultivation of algae consortium in a dairy farm wastewater for
biodiesel production. Water Resour. Indust. 2015; 10: 1-14.
Hightower M, Pierce SA. The energy challenge. Nature 2008; 452: 285-286.
Huang Q, Song S, Chen Z, Hu B, Chen J, Wang X. Biochar-based materials and their
applications in removal of organic contaminants from wastewater: state-of-the-art review.
Biochar 2019; 1: 45-73.
Hultberg M, Bodin H, Birgersson G. Impact on wastewater quality of biopellets composed of
Chlorella vulgaris and Aspergillus niger and lipid content in the harvested biomass. J.
Water Resour. Protect. 2019; 11: 831.
Hussain F, Shah SZ, Zhou W, Iqbal M. Microalgae screening under CO2 stress: Growth and
micro-nutrients removal efficiency. J. Photochem. Photobiol. B: Biol. 2017; 170: 91-98.

28
Huy M, Kumar G, Kim H-W, Kim S-H. Photoautotrophic cultivation of mixed microalgae
consortia using various organic waste streams towards remediation and resource
recovery. Bioresour. Technol. 2018; 247: 576-581.
Javed F, Aslam M, Rashid N, Shamair Z, Khan AL, Yasin M, et al. Microalgae-based biofuels,
resource recovery and wastewater treatment: A pathway towards sustainable biorefinery.
Fuel 2019; 255: 115826.
Jayakumar S, Yusoff MM, Rahim MHA, Maniam GP, Govindan N. The prospect of microalgal
biodiesel using agro-industrial and industrial wastes in Malaysia. Renew. Sust. Energ.
Rev. 2017; 72: 33-47.
Ji X, Jiang M, Zhang J, Jiang X, Zheng Z. The interactions of algae-bacteria symbiotic system
and its effects on nutrients removal from synthetic wastewater. Bioresour. Technol. 2018;
247: 44-50.
Johnson KR, Admassu W. Mixed algae cultures for low cost environmental compensation in
cultures grown for lipid production and wastewater remediation. J. Chem. Technol.
Biotechnol. 2013; 88: 992-998.
Jordaan E, Roux-van der Merwe M, Badenhorst J, Knothe G, Botha B. Evaluating the usability
of 19 effluents for heterotrophic cultivation of microalgal consortia as biodiesel
feedstock. J. Appl. Phycol. 2018; 30: 1533-1547.
Juneja A, Ceballos RM, Murthy GS. Effects of environmental factors and nutrient availability on
the biochemical composition of algae for biofuels production: a review. Energies 2013; 6:
4607-4638.
Khan A, Shahid A, Cheng H, Mahboob S, Al-Ghanim K, Bilal M, et al. Omics technologies for
microalgae-based fuels and chemicals; challenges and opportunities. Prot. Pept. Lett.
2018; 25.
Khan MK, Teng J-Z, Khan MI, Khan MO. Impact of globalization, economic factors and energy
consumption on CO2 emissions in Pakistan. Sci. Total Environ. 2019; 688: 424-436.
Khan S, Shamshad I, Waqas M, Nawab J, Ming L. Remediating industrial wastewater containing
potentially toxic elements with four freshwater algae. Ecol. Eng. 2017; 102: 536-541.
Khatoon H, Haris H, Rahman NA, Zakaria MN, Begum H, Mian S. Growth, proximate
composition and pigment production of Tetraselmis chuii cultured with aquaculture
wastewater. J. Ocean Univ. China 2018; 17: 641-646.
Koutra E, Grammatikopoulos G, Kornaros M. Selection of microalgae intended for valorization
of digestate from agro-waste mixtures. Waste Manage. 2018; 73: 123-129.
Kouzuma A, Watanabe K. Exploring the potential of algae/bacteria interactions. Curr. Opin.
Biotechnol. 2015; 33: 125-129.
Kumar A, Sharma S, Shah E, Parikh B, Patel A, Dixit G, et al. Cultivation of Ascochloris sp.
ADW007-enriched microalga in raw dairy wastewater for enhanced biomass and lipid
productivity. Int. J. Environ. Sci. Technol. 2018: 1-12.
Kumar AK, Sharma S, Patel A, Dixit G, Shah E. Comprehensive evaluation of microalgal based
dairy effluent treatment process for clean water generation and other value added
products. Int. J. Phytoremediation 2019; 21: 519-530.
Kumar D, Singh B. Phycoremediation of nutrients and valorisation of microalgal biomass: an
economic perspective. Application of Microalgae in Wastewater Treatment. Springer,
2019, pp. 1-15.
Kumar KS, Dahms H-U, Won E-J, Lee J-S, Shin K-H. Microalgae–A promising tool for heavy
metal remediation. Ecotoxicol. Environ. Saf. 2015; 113: 329-352.
Kumar P, Prajapati SK, Malik A, Vijay VK. Cultivation of native algal consortium in semi-
continuous pilot scale raceway pond for greywater treatment coupled with potential
methane production. J. Environ. Chem. Eng. 2017; 5: 5581-5587.
Kumar PS, Gunasundari E. Bioremediation of heavy metals. Bioremediation: Applications for
Environmental Protection and Management. Springer, 2018, pp. 165-195.
Lam MK, Lee KT, Mohamed AR. Current status and challenges on microalgae-based carbon
capture. Int. J. Greenhouse Gas Cont. 2012; 10: 456-469.
Lee CS, Lee S-A, Ko S-R, Oh H-M, Ahn C-Y. Effects of photoperiod on nutrient removal,
biomass production, and algal-bacterial population dynamics in lab-scale
photobioreactors treating municipal wastewater. Water Res. 2015; 68: 680-691.
Lee CS, Oh H-S, Oh H-M, Kim H-S, Ahn C-Y. Two-phase photoperiodic cultivation of algal–
bacterial consortia for high biomass production and efficient nutrient removal from
municipal wastewater. Bioresour. Technol. 2016; 200: 867-875.
Leong WH, Zaine SNA, Ho YC, Uemura Y, Lam MK, Khoo KS, et al. Impact of various
microalgal-bacterial populations on municipal wastewater bioremediation and its energy
feasibility for lipid-based biofuel production. J. Environ. Manage. 2019; 249: 109384.

30
Li D, Amoah PK, Chen B, Xue C, Hu X, Gao K, et al. Feasibility of growing Chlorella
sorokiniana on cooking cocoon wastewater for biomass production and nutrient removal.
Appl. Biochem. Biotechnol. 2019a; 188: 663-676.
Li H, Zhong Y, Lu Q, Zhang X, Wang Q, Liu H, et al. Co-cultivation of Rhodotorula glutinis
and Chlorella pyrenoidosa to improve nutrient removal and protein content by their
synergistic relationship. RSC Advances 2019b; 9: 14331-14342.
Li Y, Han F, Xu H, Mu J, Chen D, Feng B, et al. Potential lipid accumulation and growth
characteristic of the green alga Chlorella with combination cultivation mode of nitrogen
(N) and phosphorus (P). Bioresour. Technol. 2014; 174: 24-32.
Lin C-Y, Nguyen M-LT, Lay C-H. Starch-containing textile wastewater treatment for biogas and
microalgae biomass production. J. Clean. Prod. 2017; 168: 331-337.
Liu Y, Yildiz I. The effect of salinity concentration on algal biomass production and nutrient
removal from municipal wastewater by Dunaliella salina. Int. J. Energ. Res. 2018; 42:
2997-3006.
López-Serna R, García D, Bolado S, Jiménez JJ, Lai FY, Golovko O, et al. Photobioreactors
based on microalgae-bacteria and purple phototrophic bacteria consortia: A promising
technology to reduce the load of veterinary drugs from piggery wastewater. Sci. Total
Environ. 2019.
Lv J, Guo J, Feng J, Liu Q, Xie S. Effect of sulfate ions on growth and pollutants removal of
self-flocculating microalga Chlorococcum sp. GD in synthetic municipal wastewater.
Bioresour. Technol. 2017; 234: 289-296.
Magdouli S, Brar S, Blais J. Co-culture for lipid production: Advances and challenges. Biomass
Bioenergy 2016; 92: 20-30.
Makut BB, Das D, Goswami G. Production of microbial biomass feedstock via co-cultivation of
microalgae-bacteria consortium coupled with effective wastewater treatment: A
sustainable approach. Algal Res. 2019; 37: 228-239.
Malvis A, Hodaifa G, Halioui M, Seyedsalehi M, Sánchez S. Integrated process for olive oil mill
wastewater treatment and its revalorization through the generation of high added value
algal biomass. Water Res. 2019; 151: 332-342.
Mandotra S, Kumar P, Suseela M, Nayaka S, Ramteke P. Evaluation of fatty acid profile and
biodiesel properties of microalga Scenedesmus abundans under the influence of
phosphorus, pH and light intensities. Bioresour. Technol. 2016; 201: 222-229.
Mar CC, Fan Y, Li F-L, Hu G-R. Bioremediation of wastewater from edible oil refinery factory
using oleaginous microalga Desmodesmus sp. S1. Int. J. Phytoremediation 2016; 18:
1195-1201.
Marchão L, da Silva TL, Gouveia L, Reis A. Microalgae-mediated brewery wastewater
treatment: effect of dilution rate on nutrient removal rates, biomass biochemical
composition, and cell physiology. J. Appl. Phycol. 2018; 30: 1583-1595.
Marchello AE, Barreto DM, Lombardi AT. Effects of titanium dioxide nanoparticles in different
metabolic pathways in the freshwater microalga Chlorella sorokiniana
(trebouxiophyceae). Water, Air, Soil Pollut. 2018; 229: 48.
Matamoros V, Gutiérrez R, Ferrer I, García J, Bayona JM. Capability of microalgae-based
wastewater treatment systems to remove emerging organic contaminants: a pilot-scale
study. J. Hazard. Mater. 2015; 288: 34-42.
Mehar J, Shekh A, Malchira NU, Mudliar S. Potential of microalgae for integrated biomass
production utilizing CO2 and food industry wastewater. Application of Microalgae in
Wastewater Treatment. Springer, 2019, pp. 41-67.
Michelon W, Da Silva MLB, Mezzari MP, Pirolli M, Prandini JM, Soares HM. Effects of
nitrogen and phosphorus on biochemical composition of microalgae polyculture
harvested from phycoremediation of piggery wastewater digestate. Appl. Biochem.
Biotechnol. 2016; 178: 1407-1419.
Miklos DB, Remy C, Jekel M, Linden KG, Drewes JE, Hübner U. Evaluation of advanced
oxidation processes for water and wastewater treatment–a critical review. Water Res.
2018; 139: 118-131.
Mikulewicz M, Chojnacka K, Kawala B, Gredes T. Trace elements in living systems: From
beneficial to toxic effects. BioMed Res. Int. 2017; 2017.
Minhas AK, Hodgson P, Barrow CJ, Adholeya A. A review on the assessment of stress
conditions for simultaneous production of microalgal lipids and carotenoids. Front.
Microbiol. 2016; 7.

32
Miranda AF, Ramkumar N, Andriotis C, Höltkemeier T, Yasmin A, Rochfort S, et al.
Applications of microalgal biofilms for wastewater treatment and bioenergy production.
Biotechnol. Biofuel 2017; 10: 120.
Mishra A, Medhi K, Maheshwari N, Srivastava S, Thakur IS. Biofuel production and
phycoremediation by Chlorella sp. ISTLA1 isolated from landfill site. Bioresour.
Technol. 2018; 253: 121-129.
Mohammadi M, Mowla D, Esmaeilzadeh F, Ghasemi Y. Cultivation of microalgae in a power
plant wastewater for sulfate removal and biomass production: A batch study. J. Environ.
Chem. Eng. 2018; 6: 2812-2820.
Mohan SV, Dahiya S, Amulya K, Katakojwala R, Vanitha T. Can circular bioeconomy be fueled
by waste biorefineries—A closer look. Bioresour. Technol. Rep.2019a; 7: 100277.
Mohan SV, Sravan JS, Butti SK, Krishna KV, Modestra JA, Velvizhi G, et al. Microbial
electrochemical technology: emerging and sustainable platform. Microbial Electrochem.
Technol. Elsevier, 2019b, pp. 3-18.
Mohd-Salleh SNA, Mohd-Zin NS, Othman N. A review of wastewater treatment using natural
material and its potential as aid and composite coagulant. Sains Malaysiana 2019; 48:
155-164.
Molinuevo-Salces B, Riaño B, Hernández D, García-González MC. Microalgae and wastewater
treatment: Advantages and disadvantages. Microalgae biotechnology for development of
biofuel and wastewater treatment. Springer, 2019, pp. 505-533.
Morée A, Beusen A, Bouwman A, Willems W. Exploring global nitrogen and phosphorus flows
in urban wastes during the twentieth century. Global Biogeochem. Cycles 2013; 27: 836-
846.
Mousavi S, Najafpour GD, Mohammadi M, Seifi MH. Cultivation of newly isolated microalgae
Coelastrum sp. in wastewater for simultaneous CO2 fixation, lipid production and
wastewater treatment. Bioprocess Biosystems Eng. 2018; 41: 519-530.
Moussa ID-B, Athmouni K, Chtourou H, Ayadi H, Sayadi S, Dhouib A. Phycoremediation
potential, physiological, and biochemical response of Amphora subtropica and
Dunaliella sp. to nickel pollution. J. Appl. Phycol. 2018; 30: 931-941.
Mühlroth A, Winge P, El Assimi A, Jouhet J, Marechal E, Hohmann-Marriott MF, et al.
Mechanisms of phosphorus acquisition and lipid class remodelling under P limitation in a
marine microalga. Plant Physiol. 2017: pp. 00621.2017.
Nalley JO, Stockenreiter M, Litchman E. Community ecology of algal biofuels: complementarity
and trait-based approaches. Indust. Biotechnol. 2014; 10: 191-201.
Ng IS, Tan SI, Kao PH, Chang YK, Chang JS. Recent developments on genetic engineering of
microalgae for biofuels and bio‐based chemicals. Biotechnol. J. 2017.
Nordlander E, Olsson J, Thorin E, Nehrenheim E. Simulation of energy balance and carbon
dioxide emission for microalgae introduction in wastewater treatment plants. Algal Res.
2017; 24: 251-260.
O'Neill EA, Rowan NJ, Fogarty AM. Novel use of the alga Pseudokirchneriella subcapitata, as
an early-warning indicator to identify climate change ambiguity in aquatic environments
using freshwater finfish farming as a case study. Sci. Total Environ. 2019.
Odjadjare E, Mutanda T, Chen Y-F, Olaniran A. Evaluation of Pre-chlorinated wastewater
effluent for microalgal cultivation and biodiesel production. Water 2018; 10: 977.
Paliwal C, Mitra M, Bhayani K, Bharadwaj VS, Ghosh T, Dubey S, et al. Abiotic stresses as
tools for metabolites in microalgae. Bioresour. Technol. 2017.
Pancha I, Chokshi K, Mishra S. Industrial Wastewater-Based Microalgal Biorefinery: A Dual
Strategy to Remediate Waste and Produce Microalgal Bioproducts. Application of
Microalgae in Wastewater Treatment. Springer, 2019, pp. 173-193.
Patil L, Kaliwal B. Effect of CO2 concentration on growth and biochemical composition of
newly isolated indigenous microalga Scenedesmus bajacalifornicus BBKLP-07. Appl.
Biochem. Biotechnol. 2017; 182: 335-348.
Peng L, Zhang Z, Lan CQ, Basak A, Bond N, Ding X, et al. Alleviation of oxygen stress on
Neochloris oleoabundans: effects of bicarbonate and pH. J. Appl. Phycol. 2017; 29: 143-
152.
Piligaev A, Sorokina K, Shashkov M, Parmon V. Screening and comparative metabolic profiling
of high lipid content microalgae strains for application in wastewater treatment.
Bioresour. Technol. 2017.
Piotrowska-Niczyporuk A, Bajguz A, Talarek M, Bralska M, Zambrzycka E. The effect of lead
on the growth, content of primary metabolites, and antioxidant response of green alga

34
 Acutodesmus obliquus (Chlorophyceae). Environ. Sci. Pollut. Res. 2015; 22: 19112-
19123.
Prasad M, Varma A, Kumari P, Mondal P. Production of lipid-containing microalgal biomass
and simultaneous removal of nitrate and phosphate from synthetic wastewater. Environ.
Technol. 2017: 1-13.
Qin L, Wang Z, Sun Y, Shu Q, Feng P, Zhu L, et al. Microalgae consortia cultivation in dairy
wastewater to improve the potential of nutrient removal and biodiesel feedstock
production. Environ. Sci. Pollut. Res. 2016; 23: 8379-8387.
Qin L, Wei D, Wang Z, Alam MA. Advantage assessment of mixed culture of Chlorella vulgaris
and Yarrowia lipolytica for treatment of liquid digestate of yeast industry and
cogeneration of biofuel feedstock. Appl. Biochem. Biotechnol. 2018: 1-14.
Qiu Y, Zu Y, Song C, Xie M, Qi Y, Kansha Y, et al. Soybean processing wastewater purification
via Chlorella L166 and L38 with potential value-added ingredients production.
Bioresour. Technol. Rep. 2019; 7: 100195.
Qu Z, Duan P, Cao X, Liu M, Lin L, Li M. Comparison of monoculture and mixed culture
(Scenedesmus obliquus and wild algae) for C, N, and P removal and lipid production.
Environ. Sci. Pollut. Res. 2019: 1-8.
Raeisossadati M, Vadiveloo A, Bahri PA, Parlevliet D, Moheimani NR. Treating anaerobically
digested piggery effluent (ADPE) using microalgae in thin layer reactor and raceway
pond. J. Appl. Phycol. 2019: 1-9.
Ramanan R, Kim B-H, Cho D-H, Oh H-M, Kim H-S. Algae–bacteria interactions: evolution,
ecology and emerging applications. Biotechnol. Adv. 2016; 34: 14-29.
Ramlow H, Machado RAF, Marangoni C. Direct contact membrane distillation for textile
wastewater treatment: a state of the art review. Water Sc. Technol. 2017: wst2017449.
Rashid N, Park W-K, Selvaratnam T. Binary culture of microalgae as an integrated approach for
enhanced biomass and metabolites productivity, wastewater treatment, and
bioflocculation. Chemosphere 2018; 194: 67-75.
Rawat I, Kumar RR, Mutanda T, Bux F. Dual role of microalgae: phycoremediation of domestic
wastewater and biomass production for sustainable biofuels production. Appl. Energ.
2011; 88: 3411-3424.
Renuka N, Sood A, Ratha SK, Prasanna R, Ahluwalia AS. Evaluation of microalgal consortia for
treatment of primary treated sewage effluent and biomass production. J. Appl. Phycol.
2013; 25: 1529-1537.
Roostaei J, Zhang Y. Spatially Explicit Life Cycle Assessment: Opportunities and challenges of
wastewater-based algal biofuels in the United States. Algal Res. 2016.
Rugnini L, Costa G, Congestri R, Antonaroli S, di Toppi LS, Bruno L. Phosphorus and metal
removal combined with lipid production by the green microalga Desmodesmus sp.: An
integrated approach. Plant Physiol. Biochem. 2018; 125: 45-51.
Ryu B-G, Kim J, Han J-I, Yang J-W. Feasibility of using a microalgal-bacterial consortium for
treatment of toxic coke wastewater with concomitant production of microbial lipids.
Bioresour. Technol. 2017; 225: 58-66.
Sadegh H, Ali GA, Gupta VK, Makhlouf ASH, Shahryari-ghoshekandi R, Nadagouda MN, et al.
The role of nanomaterials as effective adsorbents and their applications in wastewater
treatment. J. Nanostruct. Chem. 2017: 1-14.
Sakarika M, Kornaros M. Effect of pH on growth and lipid accumulation kinetics of the
microalga Chlorella vulgaris grown heterotrophically under sulfur limitation. Bioresour.
Technol. 2016; 219: 694-701.
Sakarika M, Kornaros M. Kinetics of growth and lipids accumulation in Chlorella vulgaris
during batch heterotrophic cultivation: Effect of different nutrient limitation strategies.
Bioresour. Technol. 2017; 243: 356-365.
Salama E-S, Kurade MB, Abou-Shanab RA, El-Dalatony MM, Yang I-S, Min B, et al. Recent
progress in microalgal biomass production coupled with wastewater treatment for biofuel
generation. Renew. Sust. Energ. Rev. 2017; 79: 1189-1211.
Shah MMR. Astaxanthin production by microalgae Haematococcus pluvialis through wastewater
treatment: Waste to Resource. Application of Microalgae in Wastewater Treatment.
Springer, 2019, pp. 17-39.
Shahid A, Ishfaq M, Ahmad MS, Malik S, Farooq M, Hui Z, et al. Bioenergy potential of the
residual microalgal biomass produced in city wastewater assessed through pyrolysis,
kinetics and thermodynamics study to design algal biorefinery. Bioresour. Technol. 2019:
121701.

36
Shahid A, Khan AZ, Liu T, Malik S, Afzal I, Mehmood MA. Production and processing of algal
biomass. Algae Based Polymers, Blends and Composites. Elesvier, 2017, pp. 273.
Shahid A, Khan F, Ahmad N, Farooq M, Mehmood MA. Microalgal carbohydrates and proteins:
synthesis, extraction, applications, and challenges. In: Asraful AM, Xu J-L, Wang Z,
editors. Microalgae Biotechnology for Food, Health and High Value Products. Springer
Singapore, 2020.
Shaker S, Morowvat MH, Ghasemi Y. Effects of sulfur, iron and manganese starvation on
growth, β-carotene production and lipid profile of Dunaliella salina. J. Young Pharm.
2017; 9.
Shi K, Gao Z, Shi T-Q, Song P, Ren L-J, Huang H, et al. Reactive oxygen species-mediated
cellular stress response and lipid accumulation in oleaginous microorganisms: The state
of the art and future perspectives. Front. Microbiol. 2017; 8.
Sillanpää M, Ncibi MC, Matilainen A, Vepsäläinen M. Removal of natural organic matter in
drinking water treatment by coagulation: A comprehensive review. Chemosphere 2018;
190: 54-71.
Sooksawat N, Meetam M, Kruatrachue M, Pokethitiyook P, Inthorn D. Equilibrium and kinetic
studies on biosorption potential of charophyte biomass to remove heavy metals from
synthetic metal solution and municipal wastewater. Bioremediation J. 2016; 20: 240-251.
Srinuanpan S, Chawpraknoi A, Chantarit S, Cheirsilp B, Prasertsan P. A rapid method for
harvesting and immobilization of oleaginous microalgae using pellet-forming filamentous
fungi and the application in phytoremediation of secondary effluent. Int. J.
Phytoremediation 2018; 20: 1017-1024.
Stockenreiter M, Haupt F, Seppälä J, Tamminen T, Spilling K. Nutrient uptake and lipid yield in
diverse microalgal communities grown in wastewater. Algal Res. 2016; 15: 77-82.
Su Y, Mennerich A, Urban B. A comparison of feasible methods for microalgal biomass
determinations during tertiary wastewater treatment. Ecol. Eng. 2016; 94: 532-536.
Takeshita T, Ota S, Yamazaki T, Hirata A, Zachleder V, Kawano S. Starch and lipid
accumulation in eight strains of six Chlorella species under comparatively high light
intensity and aeration culture conditions. Bioresour. Technol. 2014; 158: 127-134.
Tang C-C, Zuo W, Tian Y, Sun N, Wang Z-W, Zhang J. Effect of aeration rate on performance
and stability of algal-bacterial symbiosis system to treat domestic wastewater in
sequencing batch reactors. Bioresour. Technol. 2016; 222: 156-164.
Tao R, Kinnunen V, Praveenkumar R, Lakaniemi A-M, Rintala JA. Comparison of Scenedesmus
acuminatus and Chlorella vulgaris cultivation in liquid digestates from anaerobic
digestion of pulp and paper industry and municipal wastewater treatment sludge. J. Appl.
Phycol. 2017; 29: 2845-2856.
Tsolcha ON, Tekerlekopoulou AG, Akratos CS, Aggelis G, Genitsaris S, Moustaka-Gouni M, et
al. Biotreatment of raisin and winery wastewaters and simultaneous biodiesel production
using a Leptolyngbya-based microbial consortium. J. Clean. Prod. 2017; 148: 185-193.
Tsolcha ON, Tekerlekopoulou AG, Akratos CS, Antonopoulou G, Aggelis G, Genitsaris S, et al.
A Leptolyngbya-based microbial consortium for agro-industrial wastewaters treatment
and biodiesel production. Environ. Sci. Pollut. Res. 2018; 25: 17957-17966.
Udaiyappan AFM, Hasan HA, Takriff MS, Abdullah SRS. A review of the potentials, challenges
and current status of microalgae biomass applications in industrial wastewater treatment.
J. Water Process. Eng. 2017; 20: 8-21.
Ummalyma SB, Gnansounou E, Sukumaran RK, Sindhu R, Pandey A, Sahoo D. Bioflocculation:
An alternative strategy for harvesting of microalgae–An overview. Bioresour. Technol.
2017; 242: 227-235.
Ummalyma SB, Pandey A, Sukumaran RK, Sahoo D. Bioremediation by microalgae: Current
and emerging trends for effluents treatments for value addition of waste streams.
Biosynthetic Technology and Environmental Challenges. Springer, 2018, pp. 355-375.
Varshney P, Beardall J, Bhattacharya S, Wangikar PP. Isolation and biochemical characterisation
of two thermophilic green algal species-Asterarcys quadricellulare and Chlorella
sorokiniana, which are tolerant to high levels of carbon dioxide and nitric oxide. Algal
Res. 2018; 30: 28-37.
Vitova M, Bisova K, Kawano S, Zachleder V. Accumulation of energy reserves in algae: from
cell cycles to biotechnological applications. Biotechnol. Adv. 2015; 33: 1204-1218.
Walls LE, Velasquez-Orta SB, Romero-Frasca E, Leary P, Noguez IY, Ledesma MTO. Non-
sterile heterotrophic cultivation of native wastewater yeast and microalgae for integrated

38
 municipal wastewater treatment and bioethanol production. Biochem. Eng. J. 2019; 151:
107319.
Wan J-K, Chu W-L, Kok Y-Y, Cheong K-W. Assessing the toxicity of copper oxide
nanoparticles and copper sulfate in a tropical Chlorella. J. Appl. Phycol. 2018: 1-13.
Wang F, Gao B, Su M, Dai C, Huang L, Zhang C. Integrated biorefinery strategy for tofu
wastewater biotransformation and biomass valorization with the filamentous microalga
Tribonema minus. Bioresour. Technol. 2019a; 292: 121938.
Wang J-H, Zhang T-Y, Dao G-H, Xu X-Q, Wang X-X, Hu H-Y. Microalgae-based advanced
municipal wastewater treatment for reuse in water bodies. Appl. Microbiol. Biotechnol.
2017a; 101: 2659-2675.
Wang M, Zhang S-C, Tang Q, Shi L-D, Tao X-M, Tian G-M. Organic degrading bacteria and
nitrifying bacteria stimulate the nutrient removal and biomass accumulation in
microalgae-based system from piggery digestate. Sci. Total Environ. 2019b: 134442.
Wang X, Gao S, Zhang Y, Zhao Y, Cao W. Performance of different microalgae-based
technologies in biogas slurry nutrient removal and biogas upgrading in response to
various initial CO2 concentration and mixed light-emitting diode light wavelength
treatments. J. Clean. Prod. 2017b; 166: 408-416.
Wang Y, Ho S-H, Cheng C-L, Nagarajan D, Guo W-Q, Lin C, et al. Nutrients and COD removal
of swine wastewater with an isolated microalgal strain Neochloris aquatica CL-M1
accumulating high carbohydrate content used for biobutanol production. Bioresour.
Technol. 2017c.
Wells ML, Potin P, Craigie JS, Raven JA, Merchant SS, Helliwell KE, et al. Algae as nutritional
and functional food sources: revisiting our understanding. J. Appl. Phycol. 2017; 29: 949-
982.
Wilbanks TJ, Fernandez S. Climate change and infrastructure, urban systems, and
vulnerabilities: Technical report for the US Department of Energy in support of the
national climate assessment: Island Press, 2014.
Wong Y, Ho Y, Ho K, Leung H, Yung K. Maximization of cell growth and lipid production of
freshwater microalga Chlorella vulgaris by enrichment technique for biodiesel
production. Environ. Sci. Pollut. Res. 2017; 24: 9089-9101.
Wu J-Y, Lay C-H, Chen C-C, Wu S-Y. Lipid accumulating microalgae cultivation in textile
wastewater: Environmental parameters optimization. J. Taiwan Inst. Chem. Eng. 2017a.
Wu K-c, Ho K-c, Tang C-c, Yau Y-h. The potential of foodwaste leachate as a phycoremediation
substrate for microalgal CO2 fixation and biodiesel production. Environ. Sci. Pollut. Res.
2018: 1-11.
Wu X, Yan C, Zheng H, Luo S, Liu Y, Li W, et al. Fixing CO2 and treating wastewater from
beer brewery using microalgae. J. Bio. Mater. Bioenerg. 2017b; 11: 101-105.
Xie P, Ho S-H, Peng J, Xu X-J, Chen C, Zhang Z-F, et al. Dual purpose microalgae-based
biorefinery for treating pharmaceuticals and personal care products (PPCPs) residues and
biodiesel production. Sci. Total Environ. 2019; 688: 253-261.
Xu J, Wang X, Sun S, Zhao Y, Hu C. Effects of influent C/N ratios and treatment technologies
on integral biogas upgrading and pollutants removal from synthetic domestic sewage.
Sci. Rep. 2017; 7: 10897.
Yadav G, Dash SK, Sen R. A biorefinery for valorization of industrial waste-water and flue gas
by microalgae for waste mitigation, carbon-dioxide sequestration and algal biomass
production. Sci. Total Environ. 2019; 688: 129-135.
Yewalkar-Kulkarni S, Gera G, Nene S, Pandare K, Kulkarni B, Kamble S. Exploiting phosphate-
starved cells of Scenedesmus sp. for the treatment of raw sewage. Indian J. Microbiol.
2017; 57: 241-249.
Ying K, Zimmerman W, Gilmour D. Effects of CO and pH on growth of the microalga
Dunaliella salina. J. Microb. Biochem. Technol. 2014; 6: 167-173.
Zabochnicka-Świątek M, Rygała A. The effect of biomass (Chlorella vulgaris, Scenedesmus
armatus) concentrations on Zn2+, Pb2+ and Cd2+ biosorption from zinc smelting
wastewater. Inżynieria i Ochrona Środowiska 2017; 20.
Zhang Z, Pang Z, Xu S, Wei T, Song L, Wang G, et al. Improved carotenoid productivity and
COD removal efficiency by co-culture of Rhodotorula glutinis and Chlorella vulgaris
using starch wastewaters as raw material. Appl. Biochem. Biotechnol. 2019: 1-13.
Zheng H, Liu M, Lu Q, Wu X, Ma Y, Cheng Y, et al. Balancing carbon/nitrogen ratio to improve
nutrients removal and algal biomass production in piggery and brewery wastewaters.
Bioresour. Technol. 2018; 249: 479-486.

40
Zhou K, Zhang Y, Jia X. Co-cultivation of fungal-microalgal strains in biogas slurry and biogas
purification under different initial CO2 concentrations. Sci. Rep. 2018; 8: 7786.
Zhou W, Min M, Hu B, Ma X, Liu Y, Wang Q, et al. Filamentous fungi assisted bio-
flocculation: a novel alternative technique for harvesting heterotrophic and autotrophic
microalgal cells. Sep. Purif. Technol. 2013; 107: 158-165.
Zhou W, Wang J, Chen P, Ji C, Kang Q, Lu B, et al. Bio-mitigation of carbon dioxide using
microalgal systems: Advances and perspectives. Renew. Sust. Energ. Rev. 2017; 76:
1163-1175.
Zhu B, Chen G, Cao X, Wei D. Molecular characterization of CO2 sequestration and assimilation
in microalgae and its biotechnological applications. Bioresour. Technol. 2017.
Zhu C-M, Zhang J-Y, Guan R, Hale L, Chen N, Li M, et al. Alternate succession of aggregate-
forming cyanobacterial genera correlated with their attached bacteria by co-pathways.
Sci. Total Environ. 2019; 688: 867-879.
Table 1. Overview of the conventional and modern wastewater treatment methods.
Table 2. Impact of wastewater derived nutrients on the production of biomass and metabolite
contents with the focus on microalgal phycoremediation and biorefinery ability.
Table 3. Exploitation of binary cultures for the wastewater treatment and production of value-
added products.

42
Fig. 1. Merits and demerits of algal biomass production and processing in algal biorefinery

Fig. 2. Road map of steps involved in strain development and process optimization

Fig. 3. Schematic representation of carbon neutral nature of microalgal cultivation and

processing

Fig. 3. Overview of microalgal biomass production and processing in an integrated microalgal

biorefinery to produce a variety of products in a multi-product algal biorefinery

Table 1. Overview of the conventional and modern wastewater treatment methods.

Remova
Treatm
l Referen
ent Type Principle Pollutant Merits Demerits
efficienc ces
process
y
Easy
operation,
cost-
Filter may
Removal effective, (Eyvaz
domesti get clog,
Micro- of solids 99% Remove et al.,
c, Poor
filtration, (>5-20 (dyes suspende 2017;
industri micro-
Ultra- mm) by and TC) d solid, Udaiyap
Filtrat al and pollutant
filtration, passing 74% alkalinity, pan et
ion pigment removal,
Reverse liquid (TN) organic al.,
rich membrane
osmosis, through 77% and 2017;
wastewa fouling,
Nano- porous (COD) inorganic Wang et
ter high
filtration membran contamin al.,
operationa
e ants, high 2017)
l cost
quality
treated
water
 Easy
Selective operation,
Industrial, Low
separatio no
Activated agricultura selectivity, (Crini et
n through 96% additional
carbon or l, domestic difficult al.,
binding (organic chemicals
Adsorpti aluminum, and Heavy maintenan 2019;
of pollutant , cost-
on zeolites, metal ce, Guo et
pollutant s) effective,
organic containing formation al.,
on high
polymers, wastewate of waste 2019)
absorbent metal
r products
surface binding
capacity
Dissociat
Identificat
ion and
Ecofriend ion of
hydrolysi
Heavy ly, lower commerci (Mohd-
s of >70 %
Chemical, metals, operation al scale-up Salleh et
coagulant COD,
electro, Textile, al cost, parameter al.,
Coagula into 90-100
natural- petroleum, efficient s, 2019;
tion positive %
material cosmetics pollutant electrode Sillanpä
ions; (Heavy
based wastewate removal, passivatio ä et al.,
reactive metals)
r energy n, higher 2018)
to
efficient maintenan
negative
ce
collides
Electroche Formatio Recalcitra Degradati
Partial
mical, n of nt organic on of (Affam
degradatio
Ultrasound- organic pollutants, 53-96% organic et al.,
Advance n,
based, pollutant industrial (COD), pollutants 2018;
oxidatio expensive,
Plasma, removing and 21-85 % , Miklos
n presence
ozonation, reactive pesticidal (TOC) effective, et al.,
of toxic
UV- oxidizing wastewate vast 2018)
products
radiation, species r applicabil

44
 photo- ity,
catalytic oxidize
odor
compoun
ds

Adsorpti
High
on,
efficiency
membra 88-92%
Depends on , energy-
ne- (COD),
membrane Pollutant Industrial saving,
filtration 91-98 % Challengi (Ejraei et
and removal wastewate increased
and (TS), ng scale- al.,
nanoparticle by serial r, organic membran
photo- 85-91 % up 2019)
s being treatment pollutants e
catalytic (Deterge
employed performa
degradat nts)
nce, less
ion
fouling
(Hybrid)
Filtration
of
wastewat
Filtratio 92 %
er Phenolic
n and (TS), 36 High
followed compound
coagula % treatment High (Enaime
Chemical by s, organic
nt- (COD), efficiency maintenan et al.,
coagulants treatment pollutants,
flocculat 81 % , Energy ce cost, 2019)
by suspended
ion (Fatty saving
coagulati solids,
(Hybrid) Matter)
ng
flocculan
t
 High
Act as efficiency
Heavy
absorbent and
metals, Eco- (Gautam
for the adsorptio
inorganic, 90-100 concern, et al.,
Nano- Magnetic, photolyti n
organic & % high cost, 2019;
material carbonated, c capacity,
emerging (various poor Sadegh
s metal-oxide degradati compatibl
pollutants, metals) recyclabili et al.,
on of e with
petrochem ty, toxicity 2017)
pollutant other
icals
s technique
s
Reductio Instability,
n or Radio- Eco tricky
Bacteria,
oxidation active friendly, recovery (Ali et
Fungi, 75-99%
of metals contamina non-toxic, of al.,
Biogeni Algae plant (Dyes),
by tion, cost- intracellul 2019;
c Nano- excreted 66-85 %
natural inorganic effective, arly Gautam
particles bioactive (Heavy
chemical and sustainabl synthesize et al.,
molecules - metals)
s-based organic e, energy d 2019)
based
nanoparti pollutant efficient nanopartic
cles les
Microbial- Wide
fuel cell, Oxidatio Recalcitra applicabil
Microbi Difficult
Microbial- n or nt matter, ity,
al scale-up
electrochem reduction industrial, productio
electro- >25-63 with cost- (Mohan
ical cell, of domestic n of
chemica % effective et al.,
Bio- pollutant and food- electricity
l (COD) & efficient 2019)
electrochem s by processing and other
Technol performan
ical respiring wastewate valuable
ogy ce
treatment, microbes r commodit
Microbial ies

46
 electrolysis

Low
removal
High
efficiency
Act as efficiency
Phenolic of raw-
Biomass catalysts , robust,
compound 65-99 biochar,
(algae, crop or economic
s, heavy % some
residues), absorbent al, large
metals, (dyes) types may (Huang
industrial to specific
Biochar dyes, , contain et al.,
by- degrade area, high
organic & >90 % toxins or 2019)
products, or porosity,
inorganic (phenols metals,
municipal remove less
contamina ) non-
waste the energy
nts ecofriendl
pollutant consumin
y
g
production
process
 Slow,
requires
Economic constant
al, high maintenan (Ahmad
Autotrophic Assimilat >90 %
bio- ce, low et al.,
& ion and Nitrates, (COD
Biologic degradabi applicabili 2019;
heterotrophi dissimilat phosphate ),
al lity, ty, Crini
c microbes ion of s, dairy 38-90 %
method efficient compromi and
(Bacteria, pollutant waste (Nitroge
eliminatio sed Lichtfou
Fungi etc) s n)
n of performan se, 2019)
pollutants ce due to
abiotic
factors
Compromi
sed
performan
High
Uptake ce due to
Industrial, pollutant
of abiotic
domestic 22-98 removal,
nutrients factors
wastewate % energy & (Molinu
for and
Algae, r, heavy (TP), cost evo-
biomass wastewate
Biomass Cyanobacte metals, 20-100 efficient, Salces et
and r
ria dyes, % (TN), eco- al.,
metabolit characteris
organic & >90% friendly, 2019)
e tics,
inorganic (COD) biorefiner
productio difficult
pollutants y- based
n harvesting
process
, large
area
requires

48
Table 2. Impact of wastewater derived nutrients on the production of biomass and metabolite
contents with the focus on microalgal phycoremediation and biorefinery ability.
Nutrient removal
Metabolites (%)
Wastewater- (%) Biomass
Microalgae Wastewater derived productivity Lipid Protein Carb

Phosphorus

Chemical
Nitrogen

demand
Oxygen
other

Total

Total
stress (mgL-1d-1)

Low pH, 64 0.2

Scenedesmus Brewery
high nutrient 88 30 71 (bio- 31.4 mgm
obliquus effluent
and sugars oil) (Phe

High 11
Tetraselmis
Dairy Nutrient, 83 100 - 42.5 - - mg
suecica
varying pH (Ch

High
Tribonema
Tofu-whey nutrients and 92.8 72 86.7 431.6 37 15.5 3
minus
COD
Synthetic High
Ecuadorian 52-
(Secondary nutrients, 67 - 0.6-1.8 28 - -
Chlorella sp. 93
effluent) alkaline

C. High organic
Olive-oil mill - 97 96.2 1.25 23 11 6
pyrenoidosa matter

Organic
C. Cooking
pollutant, 29.4 46 89 85.7 23.3 39.3 29
sorokiniana cocoon
high protein
Scenedesmus
Meat market High N and P 90 85 - 98.5 23.2 41.2 -
sp.

Soybean- 7.22
Moderate 2.8
Chlorella sp. processing 85 97 70.5 - mgL- -
nitrates mgL
(diluted) 1d-1

Ascochloris
Raw-Dairy High N and P 80 97 96 94-98 33.4 - -
sp.

Chlorella 50 -25.24
& Power plant Sulfate rich 22-32 (Sulfates) (Respectivel Reported biodiesel pote
Oocystis sp. y)

Tertiary
Dunaliella Moderate
treated 57.5 69 52 28.25 - - -
salina Nitrate and P
municipal
62.4
11.
Diluted High nitrates mg/
C. 388.2 mg
municipal & and 84.2 47 - 1524 g
sorokiniana mg/g (chl
industrial phosphate (FA
phy
ME)

Diluted Low pH,

Chlorella sp. winery & high organic 4.4x106
89.3 56.5 - 51 - -
MM3 piggery matter & cells/mL
(80:20) nutrients

Synthetic
Desmodesmus High P and 85 94
industrial >90 0.05 17.6 - -
sp. heavy metals (Ni) (Cu)
effluent

50
 Aqua-culture High carbon
C. vulgaris 76.5 92.7 75.5 187 9.07 47.5 19.
and pulp & nutrient

Palm oil mill

Chalymondom High nutrient 90
effluent 65 34 56 - - -
onas sp. and CO2 mgg-1
(POME)

Ascochloris
High
sp. Raw-dairy 78 98 95 102-207 34 - -
nutrients
(ADW007)

Dunaliella Diluted food High 37

>80 80 - 200 - -
tertiolecta leachate nutrients (SFA)

Coelastrum High
Cattle farm 80 100 42 281 50.7 - -
sp. nutrients

Domestic-
Chlorella sp. High 82 (Zn), 80 (Mn), 56
treatment 50 43.8 - -
(ISTLA) nutrients (Cu)
plant

Tetraselmis High 3.25x105

Aqua-culture 54 - - 17.6 33 1
chuii nutrients cells/mL

High amount
65 0.9
Dunaliella sp. Synthetic of heavy 93.6 (Ni) - -
pgcell− mg
metals

High 29.48
Synthetic
Tetraselmis sp nutrients and 90 - 80 124.5 mg-1d- - -
mariculture
solids 1

Parachlorella High 23.8

Agro-waste >98 59 39 62 - -
kessleri nutrients (FA)
 Municipal
Asterarcys
wastewater
quadricellular Nutrients 48 50 - 65 21.9 - 5
treatment
e
plant

Secondary
Low pH and
S. obliquus brewery 94 - 74 200 - 50 -
high nutrient
effluent

Paper and High pH and

S. acuminatus 100 >97 - 685 19.9 24.3 60
pulp nutrients

High solids
Swine
C. vulgaris and 71 54 78.7 157 17.4 58.8 9.
manure
ammonium

Acidic, high
Desmodesmus Oil-refinery COD &
- 53 82 270 22 - -
sp. (Dilute) phosphate,
greases
39 m
Sea-food High
Chlorella 94.5 68.4 - 77.7 27 - Chlo
processing nutrients
phyll

Table 3. Exploitation of binary cultures for the wastewater treatment and production of value-
added products.

Biomass Impact
Type of Cultivation Wastewater Nutrient
Consortia production metabol
consortia mode source removal (%)
(gL-1) (mgL-1

52
 Chlorella, Co-cultivation 84.7
Algal-
Klebsiella & in automated Dairy Farm (Nitrates) 90 2.87 -
bacteria
Acinetobacter bioreactor (COD)

Chlorella, Co-cultivation 93.5

Algal-
Klebsiella & in automated Synthetic (nitrates) 2.84 -
bacteria
Acinetobacter bioreactor 82 (COD)

Algal- C. vulgaris & 17.4-22 %

Co-cultivation Municipal 55-64 (COD) 1.1-1..42
bacteria activated sludge (lipid)

Chlorella, Synthetic 100 (COD &

Algal- Symbiotic 15.24-16.67
Scenedesmus & municipal PO4-3-P) 0.76
bacteria system (lipid)
activated sludge sewage 98 (NH4+-N)

C. sarokiniana, 98 (N)
Algal- Light-limiting Synthetic
Nitrosomonas & 88 (COD) 2.5 -
bacteria heterotrophic municipal
Dechloromonas 96 (P)
Desmodesmus sp.
Algal- Algae-based co- 52 (NH4+-N) 4.74
& nitrifying- Piggery -
bacteria cultivation 100 (TP) chlorophyll
bacteria

C. vulgaris & 80 (Ni)

Algal- Aerobic- Synthetic
Exiguobacterium 79 (Cu) - -
bacteria illuminiation (metal-rich)
profundum 56.4 (Cr)

Chlorella, Batch mode

Algal- 99.9 (TN)
Klebsiella & Photo- Paper industry 3.17 15 % crude
bacteria 95 (COD)
Acinetobacter bioreactor
97 (COD) 338 chlorph
Algal- Spongiochloris & Airlift
Petroleum 99 8.51 a, 2.92 gL-1
bacteria Hydrocabonoclastic bioreactor
(hydrocarbon) CO2 fixatio
Algal- Scenedesmus & Photo Coke 100 (TN)
- 32-40 FAM
bacteria aerobic-heterotroph illumination (Petroleum) 90 (phenol)

Scenedesmus 98 (TP)
Algal- Two-phase
Flavobacteria & Municipal 96 (TN) 1.8 22.6 % lipi
bacteria photoperiodic
Sphingobacteria 92 (COD)

Scenedesmus
Algal- Non-sterile 96 (nitrates) 2200
obliques & wild Municipal 2.74
fungal heterotrophic 100 (TAN) bio-ethanol
yeast

Algal- C. vulgaris & Synthetic

- 47.4 (TN) 0.65 59% lipids
fungal Aspergillus niger pharmaceutical

C. pyrenoidosa & 83 (TN)

Algal- Pilot-scale
Rhodotorula Piggery 53 (TP) 1.0 60% protei
fungal bioreactor
glutinis 85 (COD)

Chlorella vulgaris 80 (COD)

Algal- Co-culture 12.34
& Rhodotorula Starch 85 (organic 9.8
fungal fermenter carotenoids
glutinis acids)

74 (COD)
Algal- Scenedesmus & Seafood
Non-sterile 93 (TP) 2.17-6.64 600-1700 li
fungal Trichoderma ressei processing
44 (TN)

Algal- C. vulgaris & Photo- 80 (NH3-N &

Yeast industry 1.23-1.56 183 lipids
fungal Yarrowia lipolytica bioreactor COD)

CO2
68 (COD)
Algal- C. vulgaris & supplementation
Biogas slurry 61.7 (TN) 644.3 -
fungal Ganoderma lucidu (Photo-
64 (TP)
bioreactor)
Algal- C. vulgaris, Photobioreactor Biogas slurry 72-73 (TN & 0.41 Improve bi

54
fungal Ganoderma TP) by 89% CO
lucidum
Synthetic 100 (COD &
Algal- Chlorella & Symbiotic
municipal Nitrate) 0.05-0.7 15-18 % lip
algal Scenedesmus system
sewage 95 (PO4-3-P)

Algal- Wild-algae & Simulated 87 (Nitrate)

- 0.278 11.5 % lipi
algal Scenedesmus municipal 19 (P)

Anaerobically 98
Algal- Chlorella & Thin-layer 5.4 mgL-1d
digested (Ammonia) 2.96
algal Scenedesmus reactor lipid
Piggery 44 (COD)

70 (nitrate)
Algal- Leptolyngbya &
Non-aseptic Cheese whey 93 (COD) 0.9 124 algal o
algal Ochromonas
84 (P)

Algal- Whey
Mixed algal phylum Heterotrophic - 14.32 1910 lipid
algal processing

Algal- Sterile-
Mixed algal phylum Fish cannery - 14.02 1240 lipid
algal heterotrophic

93 (COD)
Algal- Leptolyngbya & Winery &
Mixotrophic 78 (TN) 1.3 13% lipids
algal Ochromonas raisin
99 (P)
45.8 % lipi
Algal- Native algal Pilot-scale 99.7 (nitrate)
Greywater 0.7 28 % prote
algal consortia (raceway pond) 99 (TP)
10% carbs
C. zofingiensis,
Algal- Photo- 91-95 (TP) 143-150
Scenedesmus & Dairy 5.1-5.4
algal autotrophic 57-62 (COD) mgL-1d-1 lip
Chlorella
Algal- Native microalgae Photo- 80-100 (all 54% protei
Livestock 1.93
algal consortia autotrophic nutrients) 31 lipids,
 10% carbs
Chlorella, 153.54
Algal- High-rate algal 29,470 Lha
Scenedesmus, Dairy farm 98 (COD) Ton
algal pond (HARP) Algal oil
Chlamydomonas ha−1yr−1

56
Highlights
 Microalgae are the most promising photoautotrophs to fix atmospheric carbon
 Requirement of huge amounts of freshwater to culture microalgae is challenging
 Alternatively, wastewater cultivation offers low-cost biomass production
 Wastewater nutrient stress can manipulate the microalgal metabolite content
 Mixed cultivation offers additional benefits of efficient wastewater treatment

58