Chemosphere 310 (2023) 136767
Contents lists available at ScienceDirect
Chemosphere
journal homepage: www.elsevier.com/locate/chemosphere
An insight into algicidal characteristics of Bacillus altitudinis G3 from
dysfunctional photosystem and overproduction of reactive oxygen species
Xiping Hou a, Yaoyao Yan a, Yuqin Wang b, Tao Jiang b, Xiaohui Zhang a, Xianzhu Dai a,
Yasuo Igarashi a, Feng Luo a, **, Caiyun Yang a, b, *
a
Chongqing Key Lab of Bio-resource Development for Bioenergy, College of Resources and Environment, Southwest University, Chongqing, 400715, China
Interdisciplinary Research Centre for Agriculture Green Development in Yangtze River Basin, College of Resources and Environment, Southwest University, Chongqing,
400715, China
b
H I G H L I G H T S
G R A P H I C A L A B S T R A C T
• B. altitudinis G3 from Dianchi Lake
bloom area is algicidal against
M. aeruginosa.
• Photosystem of M. aeruginosa as an
important attack target of G3.
• G3 decreased photosynthetic efficiency
and inhibited electron transfer.
• Dysfunctional
photosystem
results
overproduction of reactive oxygen
species.
• Reactive oxygen species contributed to
the algicidal effect of G3.
A R T I C L E I N F O
A B S T R A C T
Handling Editor: Tsair-Fuh
Cyanobacterial blooms negatively affect aquatic ecosystems and human health. Algicidal bacteria can efficiently
kill bloom-causing cyanobacteria. Bacillus altitudinis G3 isolated from Dianchi Lake shows high algicidal activity
against Microcystis aeruginosa. In this study, we investigated its algicidal characteristics including attack mode,
photosynthesis responses, and source and the contribution of reactive oxygen species (ROS). The results showed
that G3 efficiently and specifically killed M. aeruginosa mainly by releasing both thermolabile and thermostable
algicidal substances, which exhibited the highest algicidal activity (99.8%, 72 h) in bacterial mid-logarithmic
growth phase. The algicidal ratio under full-light conditions (99.5%, 60 h) was significantly higher than
under dark conditions (<20%, P < 0.001). G3 filtrate caused photosystem dysfunction by decreasing photo­
synthetic efficiency, as indicated by significantly decreased Fv/Fm and PIABS (P < 0.001) values. It also inhibited
photosynthetic electron transfer as indicated by significantly decreased rETR (P < 0.001), especially Q−A
downstream, as revealed by significantly decreased φEo and ψo, and increased Mo (P < 0.001). These results
indicated that the algicidal activity of G3 filtrate is light-dependent, and the cyanobacterial photosystem is an
important target. Cyanobacterial ROS and malondialdehyde contents greatly increased by 37.1% and 208% at
36 h, respectively. ROS levels decreased by 49.2% (9 h) when diuron (3-(3-4-dichlorophenyl)-1,1-dimethylurea)
partially blocked photosynthetic electron transport from QA to QB. Therefore, excessive ROS were produced from
Keywords:
Harmful cyanobacterial blooms
Photosynthesis
Electron transfer
ROS
* Corresponding author. Chongqing Key Laboratory, Research Center of Bioenergy and Bioremediation, College of Resources and Environment, Southwest Uni­
versity, Chongqing, 400715. China.
** Corresponding author.
E-mail addresses: van77@swu.edu.cn (F. Luo), yangcyice@163.com (C. Yang).
https://doi.org/10.1016/j.chemosphere.2022.136767
Received 12 May 2022; Received in revised form 20 September 2022; Accepted 3 October 2022
Available online 11 October 2022
0045-6535/© 2022 Elsevier Ltd. All rights reserved.
X. Hou et al.
Chemosphere 310 (2023) 136767
disrupted photosynthesis, especially the inhibited electron transport area in Q−A downstream, and caused severe
lipid peroxidation with significantly increased MDA content and oxidative stress in cyanobacteria. The ROS
scavenger N-acetyl-L-cysteine significantly decreased both cyanobacterial ROS levels (34%) and algicidal ratio
(52%, P < 0.05) at 39 h. Thus, excessive ROS production due to G3 filtrate administration significantly
contributed to its algicidal effect. G3 could be an excellent algicide to control M. aeruginosa blooms in waters
under suitable light conditions.
All authors approved the final manuscript and agreed to be respon­
sible for all aspects of the work.
substances were investigated. The effect of light on algicidal effect,
comprehensive response of photosynthetic functions, and source and
contribution of cyanobacterial ROS in the algicidal process were
revealed.
1. Introduction
Photosynthesis plays a vital role for cyanobacterial growth and
reproduction. The cyanobacterial photosystem is sensitive to environ­
mental changes, such as NaCl concentration (Yang et al., 2020) and
temperature (Pedersen and Miller, 2017). Thus, it has been widely used
as an indicator of the ecological impact of climate change (Strzepek
et al., 2019) and presence of pollutants (Wang et al., 2022; Ma et al.,
2022) in aquatic ecosystems. Some algicidal bacteria and their derived
substances can disrupt photosynthesis and damage the photosystem of
the target cyanobacteria. For instance, Pseudomonas sp. QJX-1 signifi­
cantly decreased the maximum quantum yield of photosystem II (PSII)
and the photosynthetic electron transport rate of M. aeruginosa (Qi et al.,
2021). Further, the supernatant of Raoultella sp. S1 inhibited the
expression of photosystem-related genes and decreased photosynthetic
efficiency and relative electron transport rate of M. aeruginosa (Li et al.,
2021a,b). Thus, it is necessary to elucidate the algicidal characteristics
of bacteria by investigating the changes in the photosynthetic system of
the target cyanobacteria.
Previous studies have shown that many algicidal bacteria and their
derived substances can increase ROS levels in algae (Shao et al., 2013;
Zhang et al., 2020). ROS are highly reactive and have toxic effects on
organisms by directly and nonspecifically damaging essential bio­
molecules, such as nucleic acids, proteins, and lipids (Yang et al., 2020).
For example, Deinococcus sp. Y35 increased the ROS production in toxic
algae Alexandrium tamarense within 0.5 h. Consequently, this induced
significant lipid peroxidation and destroyed the integrity of cell mem­
branes (Li et al., 2015). Photosynthesis is an important source of ROS in
algae. Few studies have systematically investigated the relationships
between algicidal effects, photosynthesis, and ROS levels/production to
elucidate the algicidal characteristics of bacteria.
In addition, environmental factors can significantly influence the
algicidal process, especially those that vary with diurnal rhythm and
seasons, such as light and temperature. For example, Shao et al. (2015)
found that the algicidal ratio of Bacillus sp. B50 filtrate against
M. aeruginosa NIES-843 at 30 ◦ C was significantly higher than that at
20 ◦ C (P < 0.05) (Shao et al., 2015). The algicidal activity of Strepto­
myces amritsarensis HG-16 against M. aeruginosa under dark conditions
(31.3%) was significantly lower than that under full light (98.1%) and
light/dark cycle (97.9%) conditions (P < 0.001) (Yu et al., 2019).
Bowmanella denitrificans S088 exhibited a much higher algicidal ratio
against Chlorella vulgaris in dark than in full-light (P = 0.004) (Jiang
et al., 2014). The sensitivities of algicidal activities of different bacteria
varied significantly with environmental conditions. Therefore, to suc­
cessfully control cyanobacterial blooms with specific bacteria, it is
necessary to investigate the influence of environmental factors on the
algicidal process.
Dianchi Lake is the largest plateau lake in China and has suffered
from cyanobacterial blooms (especially M. aeruginosa) for decades (Liu
et al., 2019). In this study, a bacterium strain, Bacillus altitudinis G3, with
strong algicidal activity against the tested M. aeruginosa, was isolated
from the bloom area of Dianchi Lake. This study aimed to reveal the
algicidal characteristics of G3. The algicidal mode, algicidal effect of G3
filtrate at different growth stages, and characteristics of algicidal
2. Materials and methods
2.1. Cyanobacteria and bacteria strains and culture conditions
Microcystis aeruginosa NIES-843 was purchased from the Microbial
Culture Collection at the National Institute for Environmental Studies
(NIES Collection, Japan) and cultured in MA medium (Wei et al., 2019).
It is a toxic and axenic cyanobacterial strain, and shows unicellular
morphology under laboratory culture conditions (Kaneko et al., 2007).
This strain was cultivated under light/dark (14 h/10 h) cycles with a
light intensity of 30 μmol photons m− 2 s− 1 at 28 ◦ C. The cyanobacterial
cultures were shaken twice daily. M. aeruginosa NIES-843 (logarithmic
growth stage) with an initial cell density of 4.0 × 106 cells/mL (OD680 =
0.148) was used in all experiments.
Thirty eight bacterial strains were isolated from the surface water in
a bloom area of Dianchi Lake (Yunnan, China) in July 2015 using the
dilution plate method with LB agar plates. Strain G3 showed the highest
algicidal activity against M. aeruginosa NIES-843 (Fig. S1). According to
similarity and phylogenetic analysis (Fig. S2) of the 16S rRNA gene
(GenBank accession number: MN559531), it belongs to Bacillus altitu­
dinis. Strain G3 used in the following experiments was cultured with 1/5
LB (28 ◦ C, 180 rpm) until the mid-logarithmic growth phase (cell density
was approximately 4 × 108 cells/mL), unless specified otherwise.
2.2. Assays of algicidal mode
To investigate the algicidal mode, the supernatant of G3 culture after
centrifugation (5800×g, 10 min) was filtered with a 0.22 μm sterile filter
to obtain cell-free filtrate. Cell pellets were washed three times with 15
mL sterile water and suspended in an equal volume of sterile water (15
mL) to obtain the washed cells. G3 culture, cell-free filtrate, washed
cells, 1/5 LB (control for the G3 culture and cell-free filtrate groups), and
sterile water (control for the washed cells group) were added (5% vol­
ume) into the M. aeruginosa cultures. Inoculation ratio of 5% was used in
all algicidal experiments, because it allows obvious algicidal effect
observed and no inhibiting effect of bacterial media on M. aeruginosa
growth in control group. On day 1, 2 and 3, 100 μL samples were taken
from each treatment and mixed with 100 μL Lugol’s iodine solution in a
centrifuge tube for subsequent cell counting. In order to eliminate the
contingency of the experiment and ensure the accuracy of the experi­
ment, three replicates were performed in all experiments.
2.3. Algicidal activity of G3 filtrate at different growth stages
Growth of G3 was measured based on the OD600. The growth curve
was drawn based on cell density, which was obtained according to the
relationship between OD600 and cell density. G3 filtrates at different
growth stages (2, 16, 22, 28, 32, 36, 42, 60, and 72 h) and 1/5 LB
(control group) were added (5% volume) into the M. aeruginosa cultures.
And on day 3, 100 μL samples were taken and mixed with 100 μL Lugol’s
iodine solution in a centrifuge tube for cell counting.
2
X. Hou et al.
Chemosphere 310 (2023) 136767
2.4. Characterization of the algicidal substances
using the following equation:
The thermal stability and molecular weight range of the algicidal
substances from G3 were analyzed. The G3 filtrate (15 mL) was main­
tained at − 20, 4, 25, 40, 60, 80 and 100 ◦ C for 2 h, respectively, and then
return to 25 ◦ C. And 60 mL G3 filtrate was centrifuged at 5000×g for 2 h
using a Macrosep 1 K Omega centrifugal filter (Pall Corporation, USA) to
collect fractions <1000 Da and >1000 Da (Zhang et al., 2014a). The
filtrates, two fractions that were treated and untreated at 100 ◦ C for 2 h,
and 1/5 LB (control group) were added (5% volume) into the
M. aeruginosa cultures. On day 3, 100 μL samples were taken and mixed
with 100 μL Lugol’s iodine solution in a centrifuge tube for cell counting.
Algicidal ratio (%) = (Dco – Dtr ) / Dco × 100%
(1)
where Dco and Dtr represent the cyanobacterial cell densities of the
control and treatment groups, respectively.
2.9.2. Chlorophyll a concentration
Chlorophyll a concentration was determined as follows (Chowdhury
et al., 2021): cells were harvested (10,000×g, 10 min) from 2 mL cya­
nobacterial culture, mixed with 1.8 mL 95% ethanol, and incubated at
75 ◦ C until cells were bleached. After centrifugation (10,000×g, 10 min),
the absorbance of the supernatant at 649, 665 and 750 nm was
measured using a UV–visible spectrophotometer (UV-2600, Japan). The
chlorophyll a concentration was calculated using the following equation
(Wintermans and de Mots, 1965):
2.5. Influence of light on the algicidal effect of G3 filtrate
M. aeruginosa exposed to G3 filtrate and 1/5 LB (control group, 5%
volume) were incubated in full light (30 μmol photons m− 2 s− 1) and dark
at 28 ◦ C. Cell density was measured at 12, 24, 36, 48 and 60 h.
Chl a (mg / L) = [(A665 − A750 ) × 13.7 − (A649 − A750 ) × 5.76] × V2 / V1
(2)
where V2 is the volume of the cyanobacterial sample, 2 mL; and V1 is the
volume of 95% alcohol added, 1.8 mL.
2.6. Physiological response of M. aeruginosa to the G3 filtrate
To investigate the influence of the G3 filtrate on cyanobacterial
physiology, chlorophyll a concentration, chlorophyll fluorescence
transients and malondialdehyde (MDA) content were determined.
M. aeruginosa (47.5 mL) was exposed to 2.5 mL of the G3 filtrate and 1/5
LB (control group, 5% volume) for 3 days. MDA and chlorophyll a
concentrations were determined at 12, 24, 36, 48 and 60 h. Chlorophyll
fluorescence transients were determined at 0, 12, 24, 36, 48 and 60 h.
2.9.3. Chlorophyll fluorescence transients of M. aeruginosa
The chlorophyll fluorescence transients were measured using a
portable PAM fluorometer AquaPen AP110-C (Brno, Czech Republic)
with the OJIP test (chlorophyll fluorescence in steps of O, J, I and P) (Ji
et al., 2018). The saturating light was 900 μmol m− 2⋅s− 1 and supplied by
red (620 nm) light-emitting diodes (LEDs). The fluorescence intensities
of the O-, J- and I-steps were recorded at 50 μs, 2 ms and 30 ms,
respectively. Maximal fluorescence intensity was recorded for the
P-step. The cyanobacterial cultures (3 mL) were subjected to dark
adaptation for 10 min before measurement. Parameters including
ABS/RC, Fv/Fm, PIABS, rETR, Mo, ψo, φEo, DIo/RC and φDo were
measured. A detailed description of the chlorophyll fluorescence pa­
rameters and the corresponding calculations are listed in Table S1
(Babaei et al., 2020; Gobler, 2020).
2.7. ROS production from photosynthesis
To verify whether photosynthesis in cyanobacteria exposed to the G3
filtrate can produce additional ROS, cyanobacteria were exposed to the
G3 filtrate and 3-(3-4-dichlorophenyl)-1, 1-dimethylurea (DCMU).
DCMU can replace the QB-binding site on the D1 protein, thereby
blocking electron transport after QA and decreasing the resultant elec­
tron flux (Liu et al., 2020). DCMU (AbMole, America) was prepared
using DMSO as a stock solution (10 mM), and a final concentration of 5
μM was used in the experiment. M. aeruginosa (47.5 mL) was exposed to:
(1) 2.5 mL G3 filtrate (5% volume); (2) 25 μL DCMU for 0.5 h, followed
by addition of G3 filtrate (2.5 mL); (3) 25 μL DCMU (control group). ROS
content and cell density were measured at 3, 6 and 9 h after the addition
of the G3 filtrate.
2.9.4. MDA content
The content of cyanobacterial MDA was detected using an MDA ac­
tivity assay kit (Geruisi, Suzhou, China). It was operated according to the
instructions of the manufacturer (Vavilin et al., 1998). In brief, cells
were harvested from 1 mL cyanobacterial culture by centrifugation
(8000×g, 10 min) and suspended in the extraction solution (1 mL).
Then, cells were broken three times in a bead beater (6.5 m/s, 60 s, MP
FastPrep-24, MP Biomedicals, USA). After centrifugation (13,000×g,
4 ◦ C, 10 min), 200 μL supernatant was mixed with 300 μL working so­
lution and incubated at 90 ◦ C for 30 min. Finally, the absorbance of the
supernatant after centrifugation (13,000×g, 4 ◦ C, 10 min) was deter­
mined at 532 and 600 nm using a microplate reader (BioTek, USA).
2.8. Influence of ROS on the algicidal effect of the G3 filtrate
The influence of ROS to the algicidal effect in cyanobacteria exposed
to G3 filtrate was investigated. Subsequently, the ROS content and
algicidal ratio in M. aeruginosa exposed to G3 filtrate with and without
N-acetyl-L-cysteine (NAC) were determined. NAC is a ROS scavenger
(Liu et al., 2020), and is used to remove partial ROS in M. aeruginosa.
NAC (Sangon Biotech, China) was dissolved in deionized water to pre­
pare a stock solution (400 mM), and a final concentration of 0.2 mM was
used in the experiment. M. aeruginosa (47.5 mL) was exposed to: (1) 2.5
mL G3 filtrate (5% volume), (2) G3 filtrate + 25 μL NAC, (3) 1/5 LB, (4)
1/5 LB + 25 μL NAC. For groups (2) and (4), NAC was added 24 h after
G3 filtrate or 1/5 LB addition. Since NAC was rapidly consumed in the
cyanobacterial culture, ROS content and cell density were determined at
9, 12 and 15 h after the addition of NAC (Wu et al., 2021).
2.9.5. ROS content
The intracellular ROS content of cyanobacteria was measured in
accordance to the manufacturer instructions by using the 2′ , 7′ dichlorofluorescein diacetate (DCFH-DA) fluorescent probe (Jiancheng,
China) (Zhang et al., 2013). In brief, cyanobacterial cells were harvested
from 1 mL cyanobacterial culture by centrifugation (8000×g, 10 min)
and incubated with DCFH-DA (5 μM) at 37 ◦ C in the dark for 20 min.
Cells were harvested and washed with MA three times and then sus­
pended in 2 mL MA. The ROS content, represented by dual-wavelength
ratio spectrometry (488/525 nm), was measured using a fluorescence
spectrometer (Hitachi, F-7000, Japan).
2.9. Analysis methods
2.9.1. Cyanobacterial cell density and the algicidal ratio
After staining with Lugol’s iodine solution in equal volumes, the
cyanobacterial cell density was determined using a hemocytometer
(Lack, 1971). The algicidal ratio based on the cell density was calculated
2.10. Statistical analysis
Statistical analysis (e.g., T-test) was performed using Excel 2016
software (Microsoft Office). One-way analysis of variance (ANOVA) was
3
X. Hou et al.
Chemosphere 310 (2023) 136767
Fig. 1. Algicidal ratios of culture, cell-free filtrate and washed cells of G3 against M. aeruginosa (a), and algicidal ratios of cell-free filtrates from different growth
stages of G3 (day 3) and growth curve of G3 (b). Different lowercase letters represent significant differences among different groups at a given time (h). (P < 0.05).
performed using SPSS Statistics 26.
h). Initially, the algicidal ability of the G3 filtrate increased with bac­
terial growth, reached its highest in the mid-logarithmic phase, and then
remained at a relatively high level. These results indicate that the release
of algicidal substances increased with bacterial growth, and most algi­
cidal substances were retained in the culture. The G3 filtrate from the
mid-logarithmic phase (cell density was approximately 4 × 108 cells/
mL) was used in subsequent experiments.
The aquatic environment safety risk of G3 was estimated by inves­
tigating the influences on the growth of common freshwater algae. G3
filtrate showed high algicidal effect on strains of M. aeruginosa, including
DCM4, NIES-843 and FACHB-905 (Table S2). However, it showed no
obvious algicidal effect on common freshwater algae, such as Synecho­
cystis sp., Phormidium sp., Nostoc punctiforme, Aphanizomenon sp., Ana­
baena flos-aquae, Chlamydomonas reinhardtii, Desmodesmus communis and
Monoraphidium sp. (Table S2). And it even increased the growth of
M. wesenbergii (Table S2). Thus, G3 has a narrow algicidal spectrum. In
general, this strain has a great potential to specifically alleviate
M. aeruginosa blooms with low threats to other common freshwater
algae.
3. Results and discussion
3.1. Algicidal mode and algicidal activity of G3 extracellular substances
at different growth stages
The algicidal ratios of G3 culture, filtrate and cell groups were 54.9
± 7.2%, 53.7 ± 4.2% and 8.5 ± 9.5% at 24 h, respectively (Fig. 1a). The
algicidal ratio of the cell group was significantly lower than that of the
culture and filtrate groups (P < 0.05). However, it greatly increased later
and reached 98.0 ± 1.3% at 72 h, and showed no significant difference
from that of the other two groups (Fig. 1a, P > 0.05). The results indi­
cated that G3 killed M. aeruginosa mainly by releasing algicidal sub­
stances. Algicidal substances were gradually produced and released into
cyanobacterial cultures later in the cell group. Thus, high algicidal ac­
tivity was observed at 72 h. However, it remains unclear whether the
direct attack mode is involved in the algicidal process. Another algicidal
bacterium was isolated from B. altitudinis and named MaI11-10. It only
showed direct attack against M. aeruginosa, with an algicidal ratio of
65% in 10 days (Yang et al., 2012). For comparison, G3 had a different
algicidal mode from that of MaI11-10 and much stronger algicidal
ability. Thus, bacterial strains from the same species can have varying
algicidal effects and characteristics.
To investigate the algicidal ability of G3 extracellular substances
from different growth stages, the growth curve of G3 and the algicidal
ratio of G3 filtrates were analyzed (Fig. 1b). The cell density of G3
remained low from 0 to 12 h (lag phase), then increased sharply until 30
h (logarithmic growth phase), remained high until 32 h (stable phase),
and decreased gradually and stabilized (decline phase). Algicidal ratios
of G3 filtrate from 2 to 16 h were low (11.4 ± 1.7 and 37.9 ± 12.0%,
respectively). It significantly increased to 99.8 ± 0.3% in the midlogarithmic phase (22 h), and then decreased to 91.3–97.3% (28–72
3. 2. characterization of algicidal substances from G3
The algicidal ratio of G3 filtrates treated at -20-80 ◦ C remained high
(84.7–94.7%), followed by a sharp and significant decrease at 100 ◦ C
(69.4 ± 2.3%, Fig. 2a). The results indicated that part of the G3 algicidal
substances was heat-intolerant. On day 3, the molecular weight fractions
<1000 Da and >1000 Da showed algicidal ratios of 64 ± 7.3 and 55.4 ±
8.9%, respectively (Fig. 2b). When they were treated at 100 ◦ C, the
algicidal ratio of the <1000 Da fraction significantly decreased to 44.1%
(P < 0.05), while that of the >1000 Da fraction did not change signifi­
cantly. This indicated that G3 produced algicidal substances with
<1000 Da and >1000 Da fractions. Further, some algicidal substances
<1000 Da are thermally unstable at 100 ◦ C, while the >1000 Da
Fig. 2. Algicidal ratios of G3 filtrate (a) and its fractions (b) treated with different temperatures against M. aeruginosa. * (P < 0.05) indicates significant differences
when compared to the room temperature (25 ◦ C) group at a given time (72 h).
4
X. Hou et al.
Chemosphere 310 (2023) 136767
Fig. 3. Algicidal ratio of G3 filtrate (a) and increase ratio of algicidal ratio in full-light when compared to dark (b). * (P < 0.05), ** (P < 0.01), *** (P < 0.001)
indicate significant differences when compared to the control group at a given time (h).
Fig. 4. Effect of the G3 filtrate on cyanobacterial chlorophyll a concentration (a) and chlorophyll fluorescence parameters including Fv/Fm (b), PIABS (c), φEo (e),
DIo/RC (f) and rETR (48 h) (d). * (P < 0.05), ** (P < 0.01), *** (P < 0.001) indicate significant differences compared to the control group at a given time (h).
fractions are thermostable. Algicidal substances from different algicidal
bacteria have distinct characteristics. The molecular weight of the
algicidal substances of Bacillus sp. LP-10 was also <1000 Da, but it was
thermally stable between -80 and 121 ◦ C (Guan et al., 2014). The algi­
cidal substances of Streptomyces malaysiensis O4-6 range from 500 to
1000 Da, and they lose their algicidal activity at temperatures ≥80 ◦ C
(Zheng et al., 2013).
Additionally, the G3 filtrate showed protease and cellulase activities
(Fig. S3). Proteins and cellulose are important components of algal cells,
and some bacteria can kill algae by releasing extracellular enzymes (Kim
et al., 2009; Paul and Pohnert, 2011). For example, Pseudoalteromonas
sp. A28 can produce an extracellular serine protease that shows algicidal
activity against the diatom Skeletonema costatum NIES-324 (Lee et al.,
2000). Aeromonas bivalvium MA2, can degrade the cell wall of Botryo­
coccus braunii UTEX572 by secreting cellulase (Muñoz et al., 2014). The
fractions >1000 Da of G3 algicidal substances are thermostable,
5
X. Hou et al.
Chemosphere 310 (2023) 136767
Fig. 5. MDA content of M. aeruginosa with the treatment of G3 filtrate (a) and ROS fluorescence intensity of M. aeruginosa in different treatments with G3 filtrate and
DCMU (b). * (P < 0.05), ** (P < 0.01), *** (P < 0.001) indicate significant differences compared to the control group. Different lowercase letters represent significant
variations among different groups at a given time (h). (P < 0.05).
whereas enzymes often have a molecular weight >10,000 Da. Hence,
the extracellular protease and cellulase may not contribute to the algi­
cidal activity of G3.
inhibited.
rETR and φEo reflect the relative transfer rate of photosynthetic
electrons and the quantum yield of the light absorbed by the RC for
electron transfer (Sun et al., 2020; Zhang et al., 2018), respectively.
Electron transfer is crucial for photosynthesis (Nugent, 1996). Both rETR
and φEo were significantly lower than those of the control group (Fig. 4d
and e), indicating that the electron transfer of PS II was inhibited by the
G3 filtrate. Mo is the rate of reduced QA (Strasser et al., 2000), which was
significantly higher than the control group from 0 h (Fig. S4b, P < 0.01).
ψo represents the electron transfer efficiency beyond QA (Sun et al.,
2020). This was significantly lower than the control group from 0 h
(Fig. S4c, P < 0.01). The decrease in φEo (Fig. 4e) and ψo (Fig. S4c) and
an increase in Mo (Fig. S4b) indicate the inhibition of electron transfer in
Q−A downstream (Ni et al., 2012). DIo/RC and φDo represent the energy
loss per RC and the quantum yield of energy dissipation, respectively (RJ
et al., 2000). Both were significantly higher than those of the control
group from 0 h (Fig. 4f and Fig. S4d). In the electron transport model of
PSII, most light energy captured by the photosynthetic system is used for
photochemical reactions, and the rest dissipates into fluorescent emis­
sion and heat (Strasserf et al., 1995). These results indicate that the G3
filtrate significantly inhibited photosynthesis in M. aeruginosa by
decreasing the energy available for photochemical reactions and
inhibiting photosynthetic electron transfer. Therefore, the energy
dissipation in the form of fluorescent emission and heat (DIo/RC and
φDo) increased.
3.3. Influences of light on the algicidal effect of G3 filtrate
In the dark, the algicidal ratio of the G3 filtrate was relatively stable
from 12 to 60 h, and it was less than 20% at 60 h (Fig. 3a). The algicidal
ratio in full-light was significantly higher than in dark conditions from
24 h (P < 0.05), and it reached 99.5 ± 0.1% at 60 h. Light significantly
enhanced the increase ratio of algicidal ratio to 1149.3 ± 87.6% at 36 h
(Fig. 3b). These results indicate that light plays a vital role in the algi­
cidal process caused by the G3 filtrate. Light is a crucial abiotic factor
that influences the photosystems of photosynthetic organisms, including
algae. Photosynthesis is responsible for synthesis of organic matter in
algae and is a vital physiological process (Yu et al., 2019). Previous
studies have found that algicidal bacteria inhibit the growth of algae by
affecting the photosystem of algae. They may reduce the photosynthetic
efficiency of the photosynthetic system (Li et al., 2015), inhibit the
electron transfer of the electron transport chain, and destroy photo­
synthetic pigments (Zhang et al., 2013, 2018). Therefore, our results
imply that the photosynthetic system might be the target of the G3
filtrate.
3.4. Photosynthetic system dysfunction
To further explore the effect of the G3 filtrate on the photosynthetic
system of cyanobacteria, the chlorophyll a concentration and chloro­
phyll fluorescence parameters were measured (Fig. 4). There were sig­
nificant differences in all parameters between the G3 filtrate and control
groups (P < 0.05) (Fig. 4). For M. aeruginosa exposed to G3 filtrate,
chlorophyll a in cells increased with time and was significantly higher
than that of the control group from 24 h (Fig. 4a, P < 0.001). This is
consistent with the increase in the ABS/RC (Fig. S4a), which was
significantly higher than that in the control group from 0 h (P < 0.01).
ABS/RC represents the effective antenna size per reaction center (RC) (Ji
et al., 2018) and absorption of light energy (Liu et al., 2021). The in­
crease in chlorophyll a would improve the light energy absorption of the
photosystem. Fv/Fm and PIABS represent the maximum light energy
conversion efficiency and performance index of PS II, respectively
(Schreiber et al., 1995). They continued to decrease in the G3 filtrate
group, and were significantly lower than that of the control group from
0 h (Fig. 4b and c, P < 0.001). This indicates that the photosynthetic
efficiency of M. aeruginosa was significantly inhibited by the G3 filtrates.
Algae with inhibited photosynthesis under stress can enhance energy
absorption by increasing light-harvesting pigments (e.g., chlorophyll a)
so they can obtain sufficient energy to support normal photosynthesis
(Liu et al., 2021). Thus, the increase in chlorophyll a and ABS/RC could
be a survival strategy for cyanobacteria when photosynthesis is
3.5. Oxidative stress in algae and ROS production from photosynthesis
In general, ROS in photosynthetic organisms are mainly produced by
photosynthesis (Guo et al., 2015). When the photosynthetic process (e.
g., photosynthetic electron transport) is disrupted, ROS production in­
creases. For example, when the electron transport chain in PSII is
blocked, a large amount of ROS can be generated (Zhang et al., 2014b).
Photosynthetic electron transport was inhibited by the G3 filtrate, as
revealed by the decrease in rETR and φEo (Fig. 3d and e). Therefore, we
speculated that the disrupted cyanobacterial photosynthesis attributed
to the G3 filtrate would generate excessive ROS. The result showed that
ROS in M. aeruginosa exposed to G3 filtrate increased by 37.1% at 36 h
compared to that in the control group. As shown in Fig. 5a, the MDA
content in M. aeruginosa increased with time when exposed to the G3
filtrate, and was significantly higher than that of the control group from
36 h (P < 0.01), when it was increased by 208%. MDA is a peroxidation
product of lipids, especially those of the cell membrane. The increase in
cellular MDA content reflects the damage of cell membrane integrity
resulting from excessive free oxygen radicals (ROS) (Li et al., 2021a,b).
Some M. aeruginosa strains can produce microcystins which are toxic to
animals and humans (Li et al., 2021a,b). If the cyanobacterial cells are
lysed in short time attributed to algicidal bacteria might bring the risk of
massive intracellular microcystins release into water (Sun et al., 2018).
6
X. Hou et al.
Chemosphere 310 (2023) 136767
Fig. 6. Reduction rates of cyanobacterial ROS (a) and algicidal ratio (b) of G3 filtrate in the presence of NAC.
For safety reasons, it is better to use G3 only in the control of
non-microcystin-producing M. aeruginosa blooms, until we know
whether this strain has the ability to degrade microcystins. Most algi­
cidal bacteria can cause significant ROS increase in algae. For example,
the ROS content of Alexandrium tamarense exposed to Deinococcus sp.
Y35 was 3.28-fold higher than that in the control (Lia et al., 2015). Cyclo
(Pro-Gly), produced by Bacillus sp. Ts-12, induced increase in ROS levels
and MDA content in P. globosa (Tan et al., 2016). The above results
indicate that the G3 filtrate caused an increase in ROS, lipid peroxida­
tion and oxidative stress in algae. For the control group, MDA content
displayed a decreasing trend, indicating that algae may gradually adapt
to the medium supplemented with 1/5 LB and grow healthier.
As shown in Fig. 5b, the ROS level of M. aeruginosa in the DCMU + G3
filtrate group decreased with time. It was significantly lower than that of
the G3 filtrate group after 3 h, and it was lower by 49.2% at 9 h (Fig. 5b).
This confirms that the G3 filtrate-induced cyanobacterial photosynthesis
produces excessive ROS. Moreover, this indicates that the electron
transport inhibiting area of the G3 filtrate should be the Q−A downstream.
Because there would be no significant difference in the ROS content
between the DCMU + G3 filtrate and G3 filtrate group if the G3 filtrate
inhibited the electron transfer upstream of QA (where DCMU did not
block the electron flux). Moreover, this deduction is consistent with that
revealed by the decrease in φEo and ψo (Fig. 3e and Fig. S4c). Thus, the
results indicate that the G3 filtrate inhibited the electron transfer
downstream of QA, and this possibly caused electron overflow of the
photosynthetic electron transport chain. It is known that blockage in
photosynthetic electron transfer often increases electron spillage. This
leads to O2 activation, followed by increased ROS generation (Liu et al.,
2020). Moreover, ROS can damage the photosystem by inhibiting the
repair of PSII, especially the synthesis and repair of the D1 protein (Zhao
et al., 2014). Thus, the feedback loop formed between the over­
production of ROS and damage to photosynthesis can be lethal to cya­
nobacteria (Liu et al., 2020).
4.Conclusion
Cyanobacterial blooms frequently occur in waters and cause various
issues to ecological and human health. Bacteria have been explored as a
method for controlling cyanobacterial blooms, and their algicidal
characteristics and mechanisms are important for their practical appli­
cations. Our study provides evidence that algicidal bacterium G3 caused
excessive ROS production due to induction of photosynthesis dysfunc­
tion, especially downstream of Q−A in the photosynthetic electron
transfer chain, and that the resulting ROS are important for
M. aeruginosa death during the algicidal process. This study validates
cyanobacterial photosystems as an important attack target of algicidal
bacteria, and the results show that proper environmental conditions (e.
g., light) must be considered when using algicidal bacteria for bloom
control. In addition, the specific algicidal substances of G3 require
further purification and identification, and the influences of G3 on
microcystin production and degradation should be investigated in the
future.
Author contributions statement
Xiping Hou performed experiments, analyzed data and drafted the
manuscript, Yaoyao Yan performed partial experiments and collected
data, Yuqin Wang performed partial experiments, Tao Jiang, Xiaohui
Zhang and Xianzhu Dai participated in the experimental design, Yasuo
Igarashi and Feng Luo provided reagents and revised the manuscript,
Caiyun Yang designed the research, analyzed data and revised the
manuscript.
Funding information
This work was financed by the Natural Science Foundation Project of
CQ CSTC (grant No: cstc2018jcyjAX0629) to Caiyun Yang, National
Natural Science Foundation of China (grant No: 31600095) to Caiyun
Yang.
3.6. Influence of ROS overproduction on the algicidal effect of G3 filtrate
Compared to the G3 filtrate group, the ROS content in the G3 filtrate
+ NAC group decreased by 2.2%, 52.0% and 49% at 33, 36 and 39 h (9,
12 and 15 h after the addition of NAC), respectively (Fig. 6a). Similar to
the ROS content, the algicidal ratio of the G3 filtrate + NAC group also
decreased as compared to the G3 filtrate group. It decreased by 14.8%,
34.0% and 28.6% at 33, 36 and 39 h, respectively (Fig. 6b). These results
suggested that NAC scavenged part of the ROS and reduced the algicidal
effect of the G3 filtrate. The decrease in ROS reduced damage of the G3
filtrate to the algae. Therefore, ROS attributed to the cyanobacterial
death during the algicidal process (Lee et al., 2018). Hence, our results
imply that the synergistic effects of damage to the photosynthetic system
and the overproduction of ROS caused cyanobacterial death.
Declaration of competing interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
Data availability
Data will be made available on request.
Acknowledgments
The authors would like to thank Dr. Zhongbo Zhou from Southwest
University, and Dr. Yi Li from Henan Normal University for sharing their
7
X. Hou et al.
Chemosphere 310 (2023) 136767
valuable opinions and comments on the drafts of this manuscript.
Measurement of chlorophyll fluorescence transients was supported by
NOMLAB providing instrument and setup that greatly was appreciated.
Nugent, J.H.A., 1996. Oxygenic photosynthesis. Electron transfer in photosystem I and
photosystem II. Eur. J. Biochem. 237, 519–531.
Paul, C., Pohnert, G., 2011. Interactions of the algicidal bacterium Kordia algicida with
diatoms: regulated protease excretion for specific algal lysis. PLoS One 6, e21032.
Pedersen, D., Miller, S.R., 2017. Photosynthetic temperature adaptation during niche
diversification of the thermophilic cyanobacterium Synechococcus A/B clade. ISME J.
11 (4), 1053–1057.
Qi, J., Song, Y., Liang, J., Bai, Y., Hu, C., Liu, H., Qu, J., 2021. Growth inhibition of
Microcystis aeruginosa by sand-filter prevalent manganese-oxidizing bacterium.
Separ. Purif. Technol. 256, 117808.
RJ, S., A, S., M, T.-M., 2000. The Fluorescence Transient as a Tool to Characterizeand
Screen Photosynthetic Samples, pp. 445–483 (Chapter 25).
Schreiber, U., Bilger, W., Neubauer, C., 1995. Chlorophyll fluorescence as a nonintrusive
indicator for rapid assessment of in vivo photosynthesis. In: Schulze, E.D.,
Caldwell, M.M. (Eds.), Ecophysiology of Photosynthesis. Springer, Berlin,
Heidelberg, pp. 49–70.
Shao, J., He, Y., Chen, A., Peng, L., Luo, S., Wu, G., Zou, H., Li, R., 2015. Interactive
effects of algicidal efficiency of Bacillus sp. B50 and bacterial community on
susceptibility of Microcystis aeruginosa with different growth rates. Int. Biodeterior.
Biodegrad. 97, 1–6.
Shao, J., Jiang, Y., Wang, Z., Peng, L., Luo, S., Gu, J., Li, R., 2013. Interactions between
algicidal bacteria and the cyanobacterium Microcystis aeruginosa: lytic characteristics
and physiological responses in the cyanobacteria. Int. J. Environ. Sci. Technol. 11,
469–476.
Strasser, R.J., Srivastava, A., Tsimilli-Michael, M., 2000. The Fluorescence Transient as a
Tool to Characterize and Screen Photosynthetic Samples. Probing Photosynthesis
Mechanisms Regulation & Adaptation, pp. 445–483 (Chapter 25).
Strasserf, R.J., Srivastava, A., Govindjee, 1995. Polyphasic chlorophyll a fluorescence
transient in plants and cyanobacteria. Photochem. Photobiol. 61, 32–42.
Strzepek, R.F., Boyd, P.W., Sunda, W.G., 2019. Photosynthetic adaptation to low iron,
light, and temperature in Southern Ocean phytoplankton. Proc. Natl. Acad. Sci. USA
116 (10), 4388–4393.
Sun, C., Xu, Y., Hu, N., Ma, J., Sun, S., Cao, W., Klobucar, G., Hu, C., Zhao, Y., 2020. To
evaluate the toxicity of atrazine on the freshwater microalgae Chlorella sp. using
sensitive indices indicated by photosynthetic parameters. Chemosphere 244,
125514.
Sun, R., Sun, P., Zhang, J., Esquivel-Elizondo, S., Wu, Y., 2018. Microorganisms-based
methods for harmful algal blooms control: a review. Bioresour. Technol. 248, 12–20.
Tan, S., Hu, X., Yin, P., Zhao, L., 2016. Photosynthetic inhibition and oxidative stress to
the toxic Phaeocystis globosa caused by a diketopiperazine isolated from products of
algicidal bacterium metabolism. J. Microbiol. 54, 364–375.
Vavilin, D.V., Ducruet, J.-M., Matorin, D.N., Venediktov, P.S., Rubin, A.B., 1998.
Membrane lipid peroxidation, cell viability and photosystem II activity in the green
alga Chlorella pyrenoidosa subjected to various stress conditions. J. Photochem.
Photobiol., B: Biology 42, 233–239.
Wang, S., Wufuer, R., Duo, J., Li, W., Pan, X., 2022. Cadmium caused different toxicity to
photosystem I and photosystem II of freshwater unicellular algae Chlorella
pyrenoidosa (Chlorophyta). Toxics 10 (7), 352.
Wei, K., Jung, S., Amano, Y., Machida, M., 2019. Control of the buoyancy of Microcystis
aeruginosa via colony formation induced by regulating extracellular polysaccharides
and cationic ions. SN Appl. Sci. 1.
Wintermans, J.F., de Mots, A., 1965. Spectrophotometric characteristics of chlorophylls a
and b and their pheophytins in ethanol. Biochim. Biophys. Acta 109, 448–453.
Wu, D., Yang, C., Zhang, X., Hou, X., Zhang, S., Dai, X., Zhang, X., Igarashi, Y., Luo, F.,
2021. Algicidal Effect of Tryptoline against Microcystis Aeruginosa: Excess Reactive
Oxygen Species Production Mediated by Photosynthesis. Science of the Total
Environment, 150719.
Yang, C., Hou, X., Wu, D., Chang, W., Zhang, X., Dai, X., Du, H., Zhang, X., Igarashi, Y.,
Luo, F., 2020. The characteristics and algicidal mechanisms of cyanobactericidal
bacteria, a review. World J. Microbiol. Biotechnol. 36, 188.
Yang, L., Wei, H., Masaharu, K., Kenichi, I., Lin, J., Tatsuo, I., Takeshi, Y., Hiroto, M.,
2012. Isolation and characterization of bacterial isolates algicidal against a harmful
bloom-forming cyanobacterium Microcystis aeruginosa. Biocontrol Sci. 17, 107–114.
Yu, Y., Zeng, Y., Li, J., Yang, C., Zhang, X., Luo, F., Dai, X., 2019. An algicidal
Streptomyces amritsarensis strain against Microcystis aeruginosa strongly inhibits
microcystin synthesis simultaneously. Sci. Total Environ. 650, 34–43.
Zhang, B., Cai, G., Wang, H., Li, D., Yang, X., An, X., Zheng, X., Tian, Y., Zheng, W.,
Zheng, T., 2014a. Streptomyces alboflavus RPS and its novel and high algicidal
activity against harmful algal bloom species Phaeocystis globosa. PLoS One 9,
e92907.
Zhang, F., Fan, Y., Zhang, D., Chen, S., Bai, X., Ma, X., Xie, Z., Xu, H., 2020. Effect and
mechanism of the algicidal bacterium Sulfitobacter porphyrae ZFX1 on the
mitigation of harmful algal blooms caused by Prorocentrum donghaiense. Environ.
Pollut. 263, 114475.
Zhang, F., Ye, Q., Chen, Q., Fan, Y., Yao, L., Ke, L., Zheng, T., Yang, K., Zhang, D., Xu, H.,
2018. Algicidal activity of novel marine bacterium Paracoccus sp. strain Y42 against
a harmful algal-bloom-causing dinoflagellate, Prorocentrum donghaiense. Appl.
Environ. Microbiol. 84, e1015–e1018.
Zhang, H., An, X., Zhou, Y., Zhang, B., Zhang, S., Li, D., Chen, Z., Li, Y., Bai, S., Lv, J.,
Zheng, W., Tian, Y., Zheng, T., 2013. Effect of oxidative stress induced by
Brevibacterium sp. BS01 on a HAB causing species–Alexandrium tamarense. PLoS One
8, e63018.
Zhang, H., Lv, J., Peng, Y., Zhang, S., An, X., Xu, H., Zhang, J., Tian, Y., Zheng, W.,
Zheng, T., 2014b. Cell death in a harmful algal bloom causing species Alexandrium
Appendix A. Supplementary data
Supplementary data to this article can be found online at https://doi.
org/10.1016/j.chemosphere.2022.136767.
References
Babaei, A., Ranglová, K., Malapascua, J.R., Torzillo, G., Shayegan, J., Silva Benavides, A.
M., Masojídek, J., 2020. Photobiochemical changes in Chlorella g120 culture during
trophic conversion (metabolic pathway shift) from heterotrophic to phototrophic
growth regime. J. Appl. Phycol. 32, 2807–2818.
Chowdhury, R.I., Basak, R., Wahid, K.A., Nugent, K., Baulch, H., 2021. A rapid approach
to measure extracted chlorophyll-a from lettuce leaves using electrical impedance
spectroscopy. Water, Air, Soil Pollut. 232.
Gobler, C.J., 2020. Climate change and harmful algal blooms: insights and perspective.
Harmful Algae 91, 101731.
Guan, C., Guo, X., Cai, G., Zhang, H., Li, Y., Zheng, W., Zheng, T., 2014. Novel algicidal
evidence of a bacterium Bacillus sp. LP-10 killing Phaeocystis globosa, a harmful
algal bloom causing species. Biol. Control 76, 79–86.
Guo, X., Liu, X., Pan, J., Yang, H., 2015. Synergistic algicidal effect and mechanism of
two diketopiperazines produced by Chryseobacterium sp. strain GLY-1106 on the
harmful bloom-forming Microcystis aeruginosa. Sci. Rep. 5, 14720.
Ji, X., Cheng, J., Gong, D., Zhao, X., Qi, Y., Su, Y., Ma, W., 2018. The effect of NaCl stress
on photosynthetic efficiency and lipid production in freshwater microalgaScenedesmus obliquus XJ002. Sci. Total Environ. 633, 593–599.
Jiang, X., Ren, C., Hu, C., Zhao, Z., 2014. Isolation and algicidal characterization of
Bowmanella denitrificans S088 against Chlorella vulgaris. World J. Microbiol.
Biotechnol. 30, 621–629.
Kaneko, T., Nakajima, N., Okamoto, S., Suzuki, I., Tanabe, Y., Tamaoki, M.,
Nakamura, Y., Kasai, F., Watanabe, A., Kawashima, K., Kishida, Y., Ono, A.,
Shimizu, Y., Takahashi, C., Minami, C., Fujishiro, T., Kohara, M., Katoh, M.,
Nakazaki, N., Nakayama, S., Yamada, M., Tabata, S., Watanabe, M.M., 2007.
Complete genomic structure of the bloom-forming toxic cyanobacterium Microcystis
aeruginosa NIES-843. DNA Res. 14 (6), 247–256.
Kim, J.D., Kim, J.Y., Park, J.K., Lee, C.G., 2009. Selective control of the Prorocentrum
minimum harmful algal blooms by a novel algal-lytic bacterium Pseudoalteromonas
haloplanktis AFMB-008041. Mar. Biotechnol. 11, 463–472.
Lack, T.J., 1971. Quantitative studies on the phytoplankton of the rivers thames and
kennet at reading. Freshw. Biol. 1, 213–224.
Lee, H.W., Park, B.S., Joo, J.H., Patidar, S.K., Choi, H.J., Jin, E., Han, M.S., 2018.
Cyanobacteria-specific algicidal mechanism of bioinspired naphthoquinone
derivative, NQ 2-0. Sci. Rep. 8, 11595.
Lee, S.-o., Kato, J., Takiguchi, N., Mitsutani, A., Kuroda, A., Ohtake, H., 2000.
Involvement of an extracellular protease in algicidal activity of the marine bacterium
Pseudoalteromonas sp. strain A28. Appl. Environ. Microbiol. 66, 4334–4339.
Li, D., Kang, X., Chu, L., Wang, Y., Song, X., Zhao, X., Cao, X., 2021a. Algicidal
mechanism of Raoultella ornithinolytica against Microcystis aeruginosa: antioxidant
response, photosynthetic system damage and microcystin degradation. Environ.
Pollut. 287, 117644.
Li, Y., Zhu, H., Lei, X., Zhang, H., Cai, G., Chen, Z., Fu, L., Xu, H., Zheng, T., 2015. The
death mechanism of the harmful algal bloom species Alexandrium tamarense induced
by algicidal bacterium Deinococcus sp. Y35. Front. Microbiol. 6, 992.
Li, D., Kang, X., Chu, L., Wang, Y., Song, X., Zhao, X., Cao, X., 2021b. Algicidal
mechanism of Raoultella ornithinolytica against Microcystis aeruginosa: antioxidant
response, photosynthetic system damage and microcystin degradation. Environ.
Pollut. 287, 117644.
Lia, Y., Zhua, H., Lei, X., Zhang, H., Guan, C., Chen, Z., Zheng, W., Xu, H., Tian, Y., Yu, Z.,
Zheng, T., 2015. The first evidence of deinoxanthin from Deinococcus sp. Y35 with
strong algicidal effect on the toxic dinoflagellate Alexandrium tamarense. J. Hazard
Mater. 290, 87–95.
Liu, J., Yang, C., Chi, Y., Wu, D., Dai, X., Zhang, X., Igarashi, Y., Luo, F., 2019. Algicidal
characterization and mechanism of Bacillus licheniformis Sp34 against Microcystis
aeruginosa in Dianchi Lake. J. Basic Microbiol. 59 (11), 1112–1124.
Liu, J., Zhang, H., Yan, L., Kerr, P.G., Zhang, S., Wu, Y., 2021. Electron transport, light
energy conversion and proteomic responses of periphyton in photosynthesis under
exposure to AgNPs. J. Hazard Mater. 401, 123809.
Liu, Q., Tang, X., Zhang, X., Yang, Y., Sun, Z., Jian, X., Zhao, Y., Zhang, X., 2020.
Evaluation of the toxic response induced by BDE-47 in a marine alga, Phaeodactylum
tricornutum, based on photosynthesis-related parameters. Aquat. Toxicol. 227,
105588.
Ma, Z., Meliana, C., Munawaroh, H.S.H., Karaman, C., Karimi-Maleh, H., Low, S.S.,
Show, P.L., 2022. Recent advances in the analytical strategies of microbial biosensor
for detection of pollutants. Chemosphere, 135515.
Muñoz, C., Hidalgo, C., Zapata, M., Jeison, D., Riquelme, C., Rivas, M., 2014. Use of
cellulolytic marine bacteria for enzymatic pretreatment in microalgal biogas
production. Appl. Environ. Microbiol. 80, 4199–4206.
Ni, L., Acharya, K., Hao, X., Li, S., Li, Y., Li, Y., 2012. Effects of artemisinin on
photosystem II performance of Microcystis aeruginosa by in vivo chlorophyll
fluorescence. Bull. Environ. Contam. Toxicol. 89, 1165–1169.
8
X. Hou et al.
Chemosphere 310 (2023) 136767
tamarense upon an algicidal bacterium induction. Appl. Microbiol. Biotechnol. 98,
7949–7958.
Zhao, L., Chen, L., Yin, P., 2014. Algicidal metabolites produced by Bacillus sp. strain B1
against Phaeocystis globosa. J. Ind. Microbiol. Biotechnol. 41, 593–599.
Zheng, X., Zhang, B., Zhang, J., Huang, L., Lin, J., Li, X., Zhou, Y., Wang, H., Yang, X.,
Su, J., Tian, Y., Zheng, T., 2013. A marine algicidal actinomycete and its active
substance against the harmful algal bloom species Phaeocystis globosa. Appl.
Microbiol. Biotechnol. 97, 9207–9215.
9