Synthesis,characterization and evaluation of antimicrobial and cytotoxic activities of biogenic silver nanoparticles synthesized from <Emphasis Type="Italic">Streptomyces xinghaiensis</Emphasis> OF1 strain

We report synthesis of silver nanoparticles (AgNPs) from Streptomyces xinghaiensis OF1 strain, which were characterised by UV–Vis and Fourier transform infrared spectroscopy, Zeta sizer, Nano tracking analyser, and Transmission electron microscopy. The antimicrobial activity of AgNPs alone, and in combination with antibiotics was evaluated against bacteria, namely Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Bacillus subtilis, and yeasts viz., Candida albicans and Malassezia furfur by using micro-dilution method. The minimum inhibitory concentration (MIC) and minimum biocidal concentration of AgNPs against bacterial and yeast strains were determined. Synergistic effect of AgNPs in combination with antibacterial and antifungal antibiotics was determined by FIC index. In addition, MTT assay was performed to study cytotoxicity of AgNPs alone and in combination with antibiotics against mouse fibroblasts and HeLa cell line. Biogenic AgNPs were stable, spherical, small, polydispersed and capped with organic compounds. The variable antimicrobial activity of AgNPs was observed against tested bacteria and yeasts. The lowest MIC (16 µg ml^?1) of AgNPs was found against P. aeruginosa, followed by C. albicans and M. furfur (both 32 µg ml^?1), B. subtilis and E. coli (both 64 µg ml^?1), and then S. aureus and Klebsiella pneumoniae (256 µg ml^?1). The high synergistic effect of antibiotics in combination with AgNPs against tested strains was found. The in vitro cytotoxicity of AgNPs against mouse fibroblasts and cancer HeLa cell lines revealed a dose dependent potential. The IC₅₀ value of AgNPs was found in concentrations of 4 and 3.8 µg ml^?1, respectively. Combination of AgNPs and antibiotics significantly decreased concentrations of both antimicrobials used and retained their high antibacterial and antifungal activity. The synthesis of AgNPs using S. xinghaiensis OF1 strain is an eco-friendly, cheap and nontoxic method. The antimicrobial activity of AgNPs could result from their small size. Remarkable synergistic effect of antibiotics and AgNPs offer their valuable potential in nanomedicine for clinical application as a combined therapy in the future.     

Growth of Ag-nanoparticles in an aqueous solution and their antimicrobial activities against Gram positive,Gram negative bacterial strains and Candida fungus

Silver nanoparticles (AgNPs) were synthesized using Ocimum sanctum (Tulsi) leaves aqueous extract as reducing as well as a capping agent in absence and presence of cetyltrimethylammonium bromide (CTAB). The resulting nanomaterials were characterized by UV–visible spectrophotometer, and transmission electron microscope. The UV–Vis spectroscopy revealed the formation of AgNPs at 400–450 nm. TEM photographs indicate that the truncated triangular silver nanoplates and/or spherical morphology of the AgNPs with an average diameter of 25 nm have been distorted markedly in presence of CTAB. The AgNPs were almost mono disperse in nature. Antimicrobial activities of AgNPs were determined by using two bacteria (Gram positive Staphylococcus aureus MTCC-3160), Gram negative Escherichia coli MTCC-450) and one species of Candida fungus (Candida albicans ATCC 90030) with Kirby-Bauer or disc diffusion method. The zone of inhibition seems extremely good showing a relatively large zone of inhibition in both Staphylococcus aureus, Escherichia coli, and Candida albicans strains.     

Exopolysaccharide-mediated silver nanoparticles produced by <Emphasis Type="Italic">Lactobacillus brevis</Emphasis> NM101-1 as antibiotic adjuvant

A green, simple and effective approach was performed to synthesize potent silver nanoparticles using bacterial exopolysaccharide as both a reducing and stabilizing agent. The formation of nanoparticles was first screened by measuring the surface plasmon resonance peak around 400 nm using UV-vis spectroscopy. The morphology of the synthesized AgNPs was determined using TEM, which indicated that the AgNPs were spherical in shape and with an average size of 11–25 nm. The presence of elemental silver of the AgNPs was confirmed by EDX analysis. The possible functional groups of EPS responsible for the reduction and stabilization of AgNPs were evaluated using FTIR. The EPS reduced AgNPs showed excellent antibacterial, and antibiofilm activities against various human pathogenic bacteria. In addition, the efficiency of AgNPs with various broad-spectrum antibiotics against the tested strains was evaluated. It is evident that, the antibacterial and antibiofilm activities of the selected antibiotics were increased in the presence of AgNPs. The increase in activity was more pronounced for gram-negative bacteria Pseudomonas aeruginosa and E. coli. Interestingly, the combination of antibiotics with AgNPs has significantly increased the membrane protein leakage and ROS generation than antibiotics or AgNPs alone. This work supports that AgNPs can be used to enhance the activity of existing antibiotics against gram-negative and gram-positive bacteria for the treatment of infectious diseases.     

Recombinant destabilase from the medicinal leech: Preparation and properties

The procedure for obtaining an active recombinant destabilase from the medicinal leech in Escherichia coli cells was developed. The plasmids encoding an analogue of native destabilase, as well as the protein forms carrying polyhistidine sequence at the Cand/or N-terminus of the polypeptide were obtained during the work. The producing strains of different forms of the protein were constructed, the cultivation process was optimized. The conditions of renaturation of destabilase recombinant forms by dialysis and using chromatographic absorbent were selected. The muramidase activity towards cell walls of Micrococcus lysodeikticus bacferia and lytic activity towards E. coli were investigated. The dependence of pH and ionic strength of the solution on the activities was determined. The total antibacterial activity of destabilase towards E. coli was shown.     

Construction of malate-sensing <Emphasis Type="Italic">Escherichia coli</Emphasis> by introduction of a novel chimeric two-component system

In an attempt to develop a high-throughput screening system for screening microorganisms which produce high amounts of malate, a MalKZ chimeric HK-based biosensor was constructed. Considering the sequence similarity among Escherichia coli (E. coli) MalK with Bacillus subtilis MalK and E. coli DcuS, the putative sensor domain of MalK was fused with the catalytic domain of EnvZ. The chimeric MalK/EnvZ TCS induced the ompC promoter through the cognate response regulator, OmpR, in response to extracellular malate. Real-time quantitative PCR and GFP fluorescence studies showed increased ompC gene expression and GFP fluorescence as malate concentration increased. By using this strategy, various chimeric TCS-based bacteria biosensors can be constructed, which may be used for the development of biochemical-producing recombinant microorganisms.     

Comparison of Antimicrobial Properties of Silver Nanoparticles Synthesized from Selected Bacteria

Green silver nanoparticle (AgNP) biosynthesis is facilitated by the enzyme mediated reduction of Ag ions by plants, fungi and bacteria. The antimicrobial activity of green AgNPs is useful to overcome the challenge of antimicrobial resistance. Antimicrobial properties of biosynthesized AgNPs depend on multiple factors including culture conditions and the microbial source. The antimicrobial activity of AgNPs biosynthesized by Pseudomonas aeruginosa ATCC 27853, Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923 and Acinetobacter baumannii (confirmed clinical isolate) were investigated in this study. Biosynthesis conditions (AgNO₃ concentration, pH, incubation temperature and incubation time) were optimized to obtain the maximum AgNP yield. Presence of AgNPs was confirmed by observing a characteristic UV–Visible absorbance peak in 420–435 nm range. AgNP biosynthesis was optimal at 0.4 g/L AgNO₃ concentration under alkaline conditions at 60–70 °C. The biosynthesized AgNPs showed higher stability compared to chemogenized AgNPs in the presence of electrolytes. AgNPs synthesized by P. aeruginosa were the most stable while NPs of S. aureus were the least stable. AgNPs synthesized by P. aeruginosa and S. aureus showed good antimicrobial potential against E. coli, P. aeruginosa, S. aureus, MRSA and Candida albicans. AgNPs synthesized by S. aureus had greater antimicrobial activity. The antimicrobial activity of NPs may vary depending on the size and the morphology of NPs.     

Evaluation of microbial globin promoters for oxygen-limited processes using <Emphasis Type="Italic">Escherichia coli</Emphasis>

Oxygen-responsive promoters can be useful for synthetic biology applications, however, information on their characteristics is still limited. Here, we characterized a group of heterologous microaerobic globin promoters in Escherichia coli. Globin promoters from Bacillus subtilis, Campylobacter jejuni, Deinococcus radiodurans, Streptomyces coelicolor, Salmonella typhi and Vitreoscilla stercoraria were used to express the FMN-binding fluorescent protein (FbFP), which is a non-oxygen dependent marker. FbFP fluorescence was monitored online in cultures at maximum oxygen transfer capacities (OTR_max) of 7 and 11 mmol L^?1 h^?1. Different FbFP fluorescence intensities were observed and the OTR_max affected the induction level and specific fluorescence emission rate (the product of the specific fluorescence intensity multiplied by the specific growth rate) of all promoters. The promoter from S. typhi displayed the highest fluorescence emission yields (the quotient of the fluorescence intensity divided by the scattered light intensity at every time-point) and rate, and together with the promoters from D. radiodurans and S. coelicolor, the highest induction ratios. These results show the potential of diverse heterologous globin promoters for oxygen-limited processes using E. coli.     

Polysaccharide-based silver nanoparticles synthesized by <Emphasis Type="Italic">Klebsiella oxytoca</Emphasis> DSM 29614 cause DNA fragmentation in <Emphasis Type="Italic">E. coli</Emphasis> cells

Silver nanoparticles (AgNPs), embedded into a specific exopolysaccharide (EPS), were produced by Klebsiella oxytoca DSM 29614 by adding AgNO₃ to the cultures during exponential growth phase. In particular, under aerobic or anaerobic conditions, two types of silver nanoparticles, named AgNPs-EPS^aer and the AgNPs-EPS^anaer, were produced respectively. The effects on bacterial cells was demonstrated by using Escherichia coli K12 and Kocuria rhizophila ATCC 9341 (ex Micrococcus luteus) as Gram-negative and Gram-positive tester strains, respectively. The best antimicrobial activity was observed for AgNPs-EPS^aer, in terms of minimum inhibitory concentrations and minimum bactericidal concentrations. Observations by transmission electron microscopy showed that the cell morphology of both tester strains changed during the exposition to AgNPs-EPS^aer. In particular, an electron-dense wrapped filament was observed in E. coli cytoplasm after 3 h of AgNPs-EPS^aer exposition, apparently due to silver accumulation in DNA, and both E. coli and K. rhizophila cells were lysed after 18 h of exposure to AgNPs-EPS^aer. The DNA breakage in E. coli cells was confirmed by the comparison of 3-D fluorescence spectra fingerprints of DNA. Finally the accumulation of silver on DNA of E. coli was confirmed directly by a significant Ag⁺ release from DNA, using the scanning electrochemical microscopy and the voltammetric determinations.     

Forage Crop <Emphasis Type="Italic">Lolium multiflorum</Emphasis> Assisted Synthesis of AgNPs and Their Bioactivities Against Poultry Pathogenic Bacteria in In Vitro

Italian ryegrass is one of main feed for livestock animals/birds. It has potential antioxidant metabolites that can improve their health and protect them against various infectious diseases. In this work, we studied synthesis of silver nanoparticles assisted by forage crop Lolium multiflorum as a green synthesis way. Potential antibacterial efficacy of these synthesized nanosized silver nanoparticles against poultry pathogenic bacteria was then studied. Aqueous extract of IRG was used as reducing agent for bio-reduction of silver salt to convert Ag⁺ to Ag⁰ metallic nano-silver. Size, shape, metallic composition, functional group, and crystalline nature of these synthesized silver nanoparticles were then characterized using UV–Vis spectrophotometer, FESEM, EDX, FT-IT, and XRD, respectively. In addition, antibacterial effects of these synthesized AgNPs against poultry pathogenic bacteria were evaluated by agar well diffusion method. UV–Vis spectra showed strong absorption peak of 440–450 nm with differ reaction time ranging from 30 min to 24 h. FESEM measurements revealed particles sizes of around 20–100 nm, majority of which were spherical in shape while a few were irregular. These biosynthesized silver nanoparticles using IRG extract exhibited strong antibacterial activities against poultry pathogenic microorganisms, including Pseudomonas aeruginosa, Salmonella typhi, Escherichia coli, and Bacillus subtilis. Overall results confirmed that IRG plant extract possessed potential bioactive compounds for converting silver ions into nanosized silver at room temperature without needing any external chemical for redox reaction. In addition, such synthesized AgNPs showed strong antibacterial activities against pathogenic bacteria responsible for infectious diseases in poultry.     

Engineering chimeric two-component system into <Emphasis Type="Italic">Escherichia coli</Emphasis> from <Emphasis Type="Italic">Paracoccus denitrificans</Emphasis> to sense methanol

Escherichia coli does not have the methanol sensing apparatus, was engineered to sense methanol by employing chimeric two-component system (TCS) strategy. A chimeric FlhS/EnvZ (FlhSZ) chimeric histidine kinase (HK) was constructed by fusing the sensing domain of Paracoccus denitrificans FlhS with the catalytic domain of E. coli EnvZ. The constructed chimeric TCS FlhSZ/OmpR could sense methanol by the expression of ompC and gfp gene regulated by ompC promoter. Real-time quantitative PCR analysis and GFP-based fluorescence analysis showed the dynamic response of the chimeric TCS to methanol. The expression of ompC and the gfp fluorescence was maximum at 0.01 and 0.5% of methanol, respectively. These results suggested that E. coli was successfully engineered to sense methanol by the introduction of chimeric HK FlhSZ. This strategy can be employed for the construction of several chimeric TCS based bacterial biosensors for the development of biochemical producing recombinant microorganisms.     

Effects of Colistin and Bacteriocins Combinations on the In Vitro Growth of <Emphasis Type="Italic">Escherichia coli</Emphasis> Strains from Swine Origin

Escherichia coli strains from swine origin, either susceptible or resistant to colistin, were grown under planktonic and biofilm cultures. After which, they were treated with antibacterial agents including nisin and enterocin DD14 bacteriocins, colistin and their combinations. Importantly, the combination of colistin, enterocin DD14 and nisin eradicated the planktonic and biofilm cultures of E. coli CIP54127 and the E. coli strains with colistin-resistance phenotype such as E. coli 184 (mcr-1 ⁺) and E. coli 289 (mcr-1 ^?), suggesting therefore that bacteriocins from lactic acid bacteria could be used as agents with antibiotic augmentation capability.     

Fabrication of silver ion exchanged zeolite using scoria and its antibacterial activity

In this study, we examined the antibacterial activity of silver ion exchanged zeolite synthesized from Cheju Scoria. We synthesized zeolite in various NaOH concentrations, but zeolite synthesized in 4 M NaOH was most similar to type A zeolite. Using the synthesized zeolite, we prepared a silver ion exchanged zeolite for studies of antibacterial activity. Antibacterial tests using agar cultures of Escherichia coli (E. coli), with silver ion exchanged zeolite showed a zone of inhibition colonies bacteria did not grow near silver ion exchanged zeolite. Furthermore, spectrophotometry demonstrated a significantly low absorbance of E. coli culture mediums when silver ion exchanged zeolite was included indicating that E. coli propagation was prevented. Through results of these experiments, we conclude that synthesized products with sodalite crystal can be synthesized from Scoria, and these are suitable to produce silver ion exchanged zeolite with antibacterial activity.     

Improved Tolerance of <Emphasis Type="Italic">Escherichia coli</Emphasis> to Propionic Acid by Overexpression of Sigma Factor RpoS

Propionic acid (PA) is an economically important compound, but large-scale microbial production of PA confronts obstacle such as acid stress on microbial cells. Here, we show that overexpressing sigma factor RpoS improves the acid tolerance of Escherichia coli. Four genes including rpoS, fur, pgi and dnaK (encoding RNA polymerase sigma factor, ferric uptake regulator, phosphoglucoisomerase, and chaperone, respectively) were independently overexpressed in E. coli. The recombinant E. coli overexpressing rpoS showed the highest PA tolerance. This strain could grow in M9 medium at pH 4.62, whereas wild type E. coli survived only at pHs above 5.12. Moreover, in the shake-flask cultivation, the E. coli strain overexpressing rpoS grew faster than wild type. Notably, the minimum inhibitory concentration of PA for this recombinant strain was 7.81 mg/mL, which was 2-fold higher in comparison with wild type. Overall these results indicated that overexpression of sigma factor rpoS significantly enhanced E. coli tolerance to PA.     

<Emphasis Type="Italic">Staphylococcus simulans</Emphasis> recombinant lysostaphin: Production,purification, and determination of antistaphylococcal activity

Staphylococcus simulans lysostaphin is an endopeptidase lysing staphylococcus cell walls by cleaving pentaglycine cross-bridges in their peptidoglycan. A synthetic gene encoding S. simulans lysostaphin was cloned in Escherichia coli cells, and producer strains were designed. The level of produced biologically active lysostaphin comprised 6-30% of total E. coli cell protein (depending on E. coli M15 or BL21 producer) under batch cultivation conditions. New methods were developed for purification of lysostaphin without affinity domains and for testing its enzymatic activity. As judged by PAGE, the purified recombinant lysostaphin is of >97% purity. The produced lysostaphin lysed cells of Staphylococcus aureus and Staphylococcus haemolyticus clinical isolates. In vitro activity and general biochemical properties of purified recombinant lysostaphin produced by M15 or BL21 E. coli strains were identical to those of recombinant lysostaphin supplied by SigmaAldrich (USA) and used as reference in other known studies. The prepared recombinant lysostaphin represents a potential product for development of enzymatic preparation for medicine and veterinary due to the simple purification scheme enabling production of the enzyme of high purity and antistaphylococcal activity.     

High-frequency underwater plasma discharge application in antibacterial activity

Plasma discharge is a novel disinfection and effectual inactivation approach to treat microorganisms in aqueous systems. Inactivation of Gram-negative Escherichia coli (E. coli) by generating high-frequency, high-voltage, oxygen (O₂) injected and hydrogen peroxide (H₂O₂) added discharge in water was achieved. The effect of H₂O₂ dose and oxygen injection rate on electrical characteristics of discharge and E. coli disinfection has been reported. Microbial log reduction dependent on H₂O₂ addition with O₂ injection was observed. The time variation of the inactivation efficiency quantified by the log reduction of the initial E. coli population on the basis of optical density measurement was reported. The analysis of emission spectrum recorded after discharge occurrence illustrated the formation of oxidant species (OH^?, H, and O). Interestingly, the results demonstrated that O₂ injected and H₂O₂ added, underwater plasma discharge had fabulous impact on the E. coli sterilization. The oxygen injection notably reduced the voltage needed for generating breakdown in flowing water and escalated the power of discharge pulses. No impact of hydrogen peroxide addition on breakdown voltage was observed. A significant role of oxidant species in bacterial inactivation also has been identified. Furthermore the E. coli survivability in plasma treated water with oxygen injection and hydrogen peroxide addition drastically reduced to zero. The time course study also showed that the retardant effect on E. coli colony multiplication in plasma treated water was favorable, observed after long time. High-frequency underwater plasma discharge based biological applications is technically relevant and would act as baseline data for the development of novel antibacterial processing strategies.     

Effects of surface charge and ligand type of Au nanoparticles on green fluorescent protein-expressing <Emphasis Type="Italic">Escherichia coli</Emphasis>

The effects of the surface charge and ligand type of three types of Au nanoparticles (NPs), namely anionic polyethylene glycol (PEG)-Au NPs, anionic citrate (Cit)-Au NPs, and cationic branched polyethylenimine (bPEI)-Au NPs, on green fluorescent protein (GFP)-expressing Escherichia coli were evaluated through the combined analysis of optical density (OD) and fluorescence intensity (FI). OD and FI can provide information about cell growth and metabolism of bacteria, respectively. The results demonstrated that PEG- and Cit-Au NPs had no major effects on the OD and FI of GFP-expressing bacteria. However, it was found that Cit-Au NPs may slightly influence cell metabolism at higher concentrations, although it is necessary to perform further in-depth study to clarify this issue. Cationic bPEI-Au NPs showed significant effects on cell density and metabolism of E. coli, with an especially strong effect on metabolism. The combined analysis of OD and FI may be useful for monitoring the effects of a wide range of nanomaterials on microorganisms.     

A homomeric geranyl diphosphate synthase-encoding gene from <Emphasis Type="Italic">Camptotheca acuminata</Emphasis> and its combinatorial optimization for production of geraniol in <Emphasis Type="Italic">Escherichia coli</Emphasis>

Geranyl diphosphate (GPP), the unique precursor for all monoterpenoids, is biosynthesized from isopentenyl diphosphate and dimethylallyl diphosphate via the head-to-tail condensation reaction catalyzed by GPP synthase (GPPS). Herein a homomeric GPPS from Camptotheca acuminata, a camptothecin-producing plant, was obtained from 5′- and 3′-rapid amplification of cDNA ends and subsequent overlap extension and convenient PCR amplifications. The truncate CaGPPS was introduced to replace ispA of pBbA5c-MevT(CO)-MBIS(CO, ispA), a de novo biosynthetic construct for farnesyl diphosphate generation, and overexpressed in Escherichia coli, together with the truncate geraniol synthase-encoding gene from C. acuminata (tCaGES), to confirm CaGPPS-catalyzed reaction in vivo. A 24.0 ± 1.3 mg L^?1 of geraniol was produced in the recombinant E. coli. The production of GPP was also validated by the direct UPLC-HRMS^E analyses. The tCaGPPS and tCaGES genes with different copy numbers were introduced into E. coli to balance their catalytic potential for high-yield geraniol production. A 1.6-fold increase of geraniol production was obtained when four copies of tCaGPPS and one copy of tCaGES were introduced into E. coli. The following fermentation conditions optimization, including removal of organic layers and addition of new n-decane, led to a 74.6 ± 6.5 mg L^?1 of geraniol production. The present study suggested that the gene copy number optimization, i.e., the ratio of tCaGPPS and tCaGES, plays an important role in geraniol production in the recombinant E. coli. The removal and addition of organic solvent are very useful for sustainable high-yield production of geraniol in the recombinant E. coli in view of that the solubility of geraniol is limited in the fermentation broth and/or n-decane.     

The Phenolic Contents and Antimicrobial Activities of Some Marine Algae from the Mediterranean Sea (Algeria)

In this study, three marine algae collected from western coast of algerian mediteranean sea (Ulva lactuca, Dictyota dichotoma, and Corallina elongata) were tested using the agar-well diffusion method for their production of antibacterial and antifungal agents on various organisms that cause diseases of humans and plants (Eschirichia coli ATCC 25922, Staphylococcus aureus ATCC 25923, Salmonella sp, Candida albicans, and Penicillium sp.). The total phenol content and antimicrobial activity were determined using different crude seaweeds extracts (methanol, diethylether, and chloroform). The results show that the chloroform extracts of (Ulva lactuca and Corallina elongata) had the highest activity against E. coli and Salmonella sp. The methanol extract obtained from (Ulva lactuca, Dictyota dichotoma, and Corallina elongata) showed antifungal activity for Candida albicans. The results of the study revealed that the seaweeds from Algeria appear to have immense potential as a source of antibacterial and antifungal compounds; they can be used in treating diseases caused by these organisms.     

Comparison of optimal conditions for mass production of carboxymethylcellulase by <Emphasis Type="Italic">Escherichia coli</Emphasis> JM109/A-68 with other recombinants in pilot-scale bioreactor

The optimal conditions for mass production of carboxymethylcellulase (CMCase) by E. coli JM109/A-68 were investigated and compared with other E. coli JM109 recombinants producing CMCase. The optimal agitation speed and aeration rate for cell growth of E. coli JM109/A- 68 were 500 rpm and 0.50 vvm in a 7 L bioreactor, whereas those for production of CMCase were 416 rpm and 0.95 vvm. The optimal vessel pressures for cell growth as well as production of CMCase in a 100 L bioreactor were 0.04 MPa. The maximal production of CMCase by E. coli JM109/A-68 under the optimized conditions in a 100 L bioreactor was 11.0 times higher than its wild type, B. velezensis A-68. Optimal conditions for mass production of CMCase by recombinants were different from those for wild strains. The higher production of CMCase by E. coli JM109/A-68 and other recombinant of E. coli seemed to result from its higher cell growth under the optimal conditions for dissolved oxygen and its mixed-growth associated production pattern compared to the growthassociated production of B. velezensis A-68.     

Inducing flocculation of non-floc-forming <Emphasis Type="Italic">Escherichia coli</Emphasis> cells

The present article reviews several approaches for inducing flocculation of Escherichia coli cells. The common industrially used bacterium E. coli does not naturally have floc-forming ability. However, there are several approaches to induce flocculation of E. coli cells. One is induction by flocculants—polyvalent inorganic salts, synthetic polymeric flocculants, or bio-based polymeric materials, including polysaccharide derivatives. Another method is the induction of spontaneous flocculation by changing the phenotypes of E. coli cells; several studies have shown that physical treatment or gene modification can endow E. coli cells with floc-forming ability. Coculturing E. coli with other microbes is another approach to induce E. coli flocculation. These approaches have particular advantages and disadvantages, and remain open to clarification of the flocculation mechanisms and improvement of the induction processes. In this review, several approaches to the induction of E. coli flocculation are summarized and discussed. This review will be a useful guide for the future development of methods for the flocculation of non-floc-forming microorganisms.