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Polymers 205 (2019) 8–26

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Carbohydrate Polymers

journal homepage: www.elsevier.com/locate/carbpol

Extremophilic exopolysaccharides: A review and new perspectives on engineering strategies and applications T ⁎ ⁎ Jia Wanga,d, David R. Salema,b,c, , Rajesh K. Sania,c,d, a Department of Chemical and Biological Engineering, South Dakota School of Mines and Technology, Rapid City, SD 57701, USA b Department of Materials and Metallurgical Engineering, South Dakota School of Mines and Technology, Rapid City, SD 57701, USA c Composite and Nanocomposite Advanced Manufacturing – Biomaterials Center (CNAM-Bio Center), Rapid City, SD 57701, USA d BuG ReMeDEE consortium, South Dakota School of Mines and Technology, Rapid City, SD 57701, USA

ARTICLE INFO ABSTRACT

Keywords: Numerous microorganisms inhabiting harsh niches produce exopolysaccharides as a significant strategy to Extremophile survive in extreme conditions. The exopolysaccharides synthesized by extremophiles possess distinctive char- Exopolysaccharide acteristics due to the varied harsh environments which stimulate the microorganisms to produce these biopo- Exopolysaccharide property lymers. Despite many bioprocesses have been designed to yield exopolysaccharides, the production of exopo- Exopolysaccharide biosynthesis lysaccharides by extremophiles is inefficient compared with mesophilic and neutrophilic exopolysaccharide Exopolysaccharide application producers. Meanwhile, the industrial development of novel extremophilic exopolysaccharides remains con- strained due to the lack of exploration. In this review, we summarize the structure and properties of various exopolysaccharides produced by extremophiles, and also discuss potential metabolic and genetic engineering strategies for enhanced yield and modified structure of extremophilic exopolysaccharides. Special focus is given to the applications of extremophilic exopolysaccharides in the areas of biomedicine, food industry, and bio- materials via nano-techniques, casting and electrospinning.

1. Introduction studied during the past several decades and applied in a variety of in- dustrial areas. In addition to , and gellan gum are In the past few decades, extremophilic microorganisms and some of currently being used in the food industry (Donot, Fontana, Baccou, & their metabolites were reported in light of their particular biosynthetic Schorr-Galindo, 2012; Rehm, 2010). Bacterial possess a mechanisms, functions, and properties which can permit the strains to great diversity of properties that may not be found in more traditional be habitant in extreme niches. Among all the products from ex- polymers of plant origin. Several EPSs have also demonstrated them- tremophiles, exopolysaccharides (EPSs) have led to significant interest selves as useful materials without the environmental disadvantages due to the increasing demand for natural polymers in various industrial associated with synthetic polymers (Chawla, Bajaj, Survase, & Singhal, fields. EPSs are high molecular weight carbohydrate biopolymers, 2009; Freitas, Alves, & Reis, 2011; Guezennec, 2002). composed of residues, and are secreted by microorganisms into Currently, it is widely accepted that extremophilic microorganisms the surrounding environment, providing certain properties and func- will provide a valuable resource for exploitation in novel biotechnolo- tions useful to the microorganisms (Nicolaus, Kambourova, & Oner, gical processes, including synthesis of unique EPSs (Bhalla, Bansal, 2010; Poli, Anzelmo, & Nicolaus, 2010). The EPS molecular chains have Kumar, Bischoff, & Sani, 2013; Nicolaus et al., 2010). The environments a broad range of molecular weights, and different microorganisms can that extremophiles inhabit are obviously more inhospitable than the synthesize a wide variety of EPSs with a diverse range of functions, such environmental pressures inducing common mesophilic and neutrophilic as intercellular signal transduction, molecular recognition, protection microbes to secrete their EPSs. Extremophiles have to adapt to hostile against predation, adhesion, biofilm formation, construction of a com- environments through unique mechanisms, and the biosynthesis of fortable extracellular environment, and pathogenic processes (Moriello EPSs is one of their vital survival mechanisms. Extremophilic micro- et al., 2003; Nicolaus et al., 1999). Some of the EPSs with valuable organisms inhabiting different extreme environments have been re- physicochemical properties have already been utilized in industry. For cognized as promising producers of EPSs, and the examination of EPS instance, among all the reported EPSs, xanthan gum has been most production by extremophiles (thermophiles, halophiles, alkaliphiles,

⁎ Corresponding authors at: Department of Chemical and Biological Engineering, South Dakota School of Mines and Technology, Rapid City, SD 57701, USA. E-mail addresses: [email protected] (D.R. Salem), [email protected] (R.K. Sani). https://doi.org/10.1016/j.carbpol.2018.10.011 Received 2 August 2018; Received in revised form 20 September 2018; Accepted 4 October 2018 Available online 09 October 2018 0144-8617/ © 2018 Elsevier Ltd. All rights reserved. J. Wang et al. Carbohydrate Polymers 205 (2019) 8–26

Fig. 1. The EPS from different kinds of extremophiles and potential applications. psychrophiles, and acidophiles) has revealed an abundance of novel microorganisms can survive in high temperatures, and their EPS pro- properties that may have strong potential in industrial applications duction has been a proposed adaptation mechanism to enable their (Fig. 1). survival in these extreme conditions. Although more and more novel extremophiles have been isolated, The growth media for thermophiles, containing as carbon and their unique EPSs characterized, the research depth of ex- and energy sources, have always been considered a primary target to be tremophilic EPSs is still not comparable with EPSs from mesophilic or optimized for maximum production of EPSs. such as neutrophilic microorganisms with regard to biosynthetic pathways, , and are the optimized carbon source for most regulatory mechanisms, and engineering strategies. It is necessary to of thermophilic bacteria for EPS production. Besides chemical compo- make a comprehensive summarization concerning the structures and sition and molecular weight, thermophilic EPSs have been character- characteristics of the recently described extremophilic EPSs, which can ized mostly in terms of thermostability. The highest decomposition provide crucial fundamentals for further exploitation of engineering temperature of 280 °C is from an EPS produced by Geobacillus tepida- strategies to obtain tailor-made extremophilic EPSs with desired yield mans (Table 1). The summarized data suggest that the type of sugar and functions. The targeted cultivation of extremophilic bacteria subunits present in the EPS may affect their thermostability. The through metabolic and genetic engineering will eventually pave the modification of monomer sugars or some other residues in EPSs can be way for industrial level applications of extremophilic EPSs. utilized to find out the active sites for certain functions (e.g., thermo- This article reviews the EPSs produced by various kinds of ex- stability) of EPSs. Although a relatively unexplored area with a sparse tremophilic bacteria, including an inventory of extremophilic EPSs of database, there is already significant evidence that EPSs from thermo- industrial interest, as well as promising engineering strategies for philes possess a broad range of interesting properties for industrial higher yield or modified molecular structure of extremophilic EPSs. applications (Nicolaus et al., 2004, 2010). The literature to date in- Moreover, the recent advances in the actual and potential applications dicates that further screening and systematic investigation of EPSs of EPSs produced by extremophilic bacteria are presented. produced by thermophiles, in conjunction with advances in under- standing the biochemistry of microbial EPS synthesis, will result in the 2. EPSs produced by different extremophile types discovery of novel biopolymers of commercial importance.

2.1. EPSs produced by thermophiles 2.2. EPSs produced by psychrophiles

Elevated temperature generally increases the rate of most chemical Psychrophiles can be isolated from Antarctic, Arctic, or deep-sea reactions and improves cumulative production in a given time frame. sediment, and they predominate in marine ecosystems (Ewert & Thus, thermophiles can be of commercial value in the synthesis of Deming, 2013; Li, Zhou, Zhang, Wang, & Zhu, 2008; Nevot, Deroncele, important compounds, and are of growing interest to many sectors of Montes, & Mercade, 2008; Nichols, Bowman, & Guezennec, 2005). The industry. Although EPS production is lower than most of the meso- EPSs from psychrophilic marine bacteria are generally carboxylated philes, the uncommonly short fermentation process, which is usually no polysaccharides, and the carboxyl groups confer a net negative charge more than 24 h, makes thermophiles important contenders as com- and acidic properties to the EPSs at the pH of seawater (pH around 8) mercially competitive EPS producers (Kambourova et al., 2009; (Caruso et al., 2017; Casillo, Parrilli et al., 2017). The negative charge Radchenkova et al., 2013; Yildiz et al., 2014). The thermophilic strains of psychrophilic EPSs can also be attributed to the phosphate groups can also typically minimize environmental contamination from meso- (Corsaro et al., 2004; Llamas et al., 2010). In the marine environment, philic microbial growth, reduce operational maintenance cost, and bacterial EPSs are essential in the production of aggregates, adhesion to improve the efficiency of substrate utilization. surface, biofilm formation and sequestering of nutrients, and provide Marine hot springs, terrestrial hot springs, and deep sea thermal protection and ecosystem stability. Due to their polyanionic property, vents have been demonstrated as the primary habitats that promote psychrophilic EPSs can accumulate cations such as metal ions, and thermophilic microbial organisms, and the majority of EPSs produced metal binding offers a potential ecological role for these biopolymers. by thermophiles have been located in these types of environments. Extracellular polysaccharides strengthen the psychrophiles’ ability to Several thermophilic bacteria in hot marine shallow vents or marine hot compete and survive in changing environmental conditions by altering springs have been shown to produce large amounts of EPSs (Manca the physical and biogeochemical micro-environment around the cells et al., 1996; Moriello et al., 2003; Nicolaus et al., 2002; Nicolaus, (Nichols, Bowman et al., 2005). The EPSs of psychrophiles in a cold Moriello, Lama, Poli, & Gambacorta, 2004). These environments are marine environment should possess the capability to protect the mi- typically characterized by their high temperature, high pressure, and croorganisms from not only the low temperature but also the relatively toxic, high inorganic or metal concentrations. Thermophilic high salinity (Caruso et al., 2018). Therefore, the EPSs secreted by the

9 .Wn tal. et Wang J. Table 1 EPSs from extremophilic bacteria.

Extremophiles Sugar carbon composition and linkage pattern Molecular Properties and activities Reference (specific conditions) source weight

Thermophiles Geobacillus thermodenitri-ficans • Monosaccharide analysis: /// 500 kDa Not tested Panosyan, Di Donato, Poli, and ArzA-6 (65 °C, pH 7.0) fructose/ (1/0.13/0.1/0.06/0.05, by relative ratio) Nicolaus, (2018) Geobacillus toebii ArzA-8 (65 °C, Fructose • Monosaccharide analysis: Mannose/galactose/glucose/ 600 kDa Not tested Panosyan, Di Donato, Poli, and pH 7.0) Arabinose (1/0.5/0.2/0.05, by relative ratio) Nicolaus, (2018) Rhodothermus marinus DSM4252T Lactose • Monosaccharide analysis: Glucose/arabinose/ (1/1.57/ 73.5 kDa Not tested Sardari et al. (2017) (65 °C, pH 7.2) 3.72, by relative ratio) Rhodothermus marinus MAT493 Maltose • Monosaccharide analysis: Glucose/arabinose/xylose/mannose 85.5 kDa Not tested Sardari et al. (2017) (65 °C, pH 7.2) (1/3.75/3.02/1.87, by relative ratio) Geobacillus sp. TS3-9 (55 °C, pH Lactose • Monosaccharide analysis: Mannose/glucose/ (1/0.14/ 3200 kDa Antioxidant activity Wang et al. (2017) 8.0) 0.06, by relative proportion) Antitumor activity Aeribacillus pallidus 418 (55 °C, pH Maltose EPS 1 EPS 1 Degradation temperature Radchenkova et al. (2013), 7.0) • Monosaccharide analysis: Mannose//galactosamine/ 700 kDa EPS1 176 °C, EPS2 226 °C Radchenkova et al. (2014), glucosamine/galactose/glucose/ (69.3/7.8/6.3/5.4/4.7/ EPS 2 Pseudoplastic rheological property Radchenkova et al. (2015) 3.4/2.9, by molar ratio) Above 1000 Foaming ability EPS 2 kDa Emulsifying activity • Monosaccharide analysis: Mannose/galactose/glucose/ galactosamine/glucosamine/ribose/arabinose (33.9/17.9/15.5/ 11.7/8.1/5.3/4.9, by molar ratio) Brevibacillus thermoruber 423 Maltose • Monosaccharide analysis: Glucose/galactose/mannose/ Not tested Biocompatibility Yildiz et al. (2014) (55 °C, pH 6.5) galactosamine/mannosamine (57.7/16.3/9.2/14.2/2.4, by percentage of abundance) Anoxybacillus sp. R4-33 (55 °C, pH Glucose • Monosaccharide analysis: Mannose/glucose (1/0.45, by relative Above 1000 Biosorption of heavy metals Zhao et al. (2014)

10 8.0) proportion) kDa Aeribacillus pallidus YM-1 (55 °C, Glucose • Monosaccharide analysis: Glucose/altrose/mannose/galactose 540 kDa Emulsifying activity Zheng et al. (2012) pH 7.5) (36.6/30.9/24.4/8.1, by molar ratio percentage) Thermus aquaticus YT-1 (60 °C, pH Not added • Monosaccharide analysis: Galactose/N-acetylgalactosamine (1/ 500 kDa Immunoregulatory activity Lin et al. (2011) 7.5) 1, by molar ratio) • Saccharide repeating unit: unit Geobacillus thermodenitri-ficans B3- Sucrose EPS 1 EPS 2 Degradation temperature 240 °C Arena et al. (2009), 72 (65 °C, pH 7.0) • Monosaccharide analysis: Glucose/mannose (1/0.3, by relative 400 kDa Hinder HSV-2 replication in human peripheral blood Nicolaus et al. (2000) ratio) mononuclear cells and partially restore the EPS 2 immunological disorders determined by HSV-2 • Monosaccharide analysis: Mannose/glucose (1/0.2, by relative ratio) Geobacillus tepidamans V264 Maltose • Monosaccharide analysis: Glucose/galactose//fructose Above 1000 Degradation temperature 280 °C Kambourova et al. (2009) (60 °C, pH 7.0) (1/0.07/0.04/0.02, by molar ratio) kDa Anti-cytotoxicity Geobacillus sp. 4004 (60 °C, pH Sucrose EPS 1 EPS 3 Not tested Moriello et al. (2003) 7.0) • Monosaccharide analysis: Glucose/mannose/galactose (1.0/0.5/ 1000 kDa 0.3, by relative ratio) EPS 2 • Monosaccharide analysis: Mannose/glucose/galactose (1.0/0.3/ Carbohydrate Polymers205(2019)8–26 trace, by relative ratio) EPS 3 • Monosaccharide analysis: Galactose/mannose/glucosamine/ arabinose (1.0/0.8/0.4/0.2, by relative ratio) • Saccharide repeating unit: Pentasaccharide unit Bacillus thermantarcti-cus (65 °C, Mannose EPS 1 EPS 2 Not tested Manca et al. (1996) pH 6.0) • Monosaccharide analysis: Mannose/glucose (1.0/0.7, by relative 300 kDa molar proportion) EPS 2 • Monosaccharide analysis: Mannose • Configuration: α-manno

(continued on next page) .Wn tal. et Wang J. Table 1 (continued)

Extremophiles Sugar carbon Monosaccharide composition and linkage pattern Molecular Properties and activities Reference (specific conditions) source weight

Psychrophiles Pseudoalteromo-nas sp. MER144 Sucrose • Monosaccharide analysis: Glucose/mannose/glucosamine/ 250 kDa Heavy metal chelation Caruso et al. (2018) (4 °C, pH 7.0) arabinose/glucuronic acid/galacturonic acid/galactose (1/0.36/ Cryoprotective activity 0.26/0.06/0.06/0.05/0.03, by relative molar ratio) Lactobacillus sakei TMW 1.411 Sucrose • Monosaccharide analysis: Glucose 3×105 kDa Not tested Prechtl et al. (2018) (10 °C, pH 5.6) Winogradskyella sp. CAL384 (4 °C, Glucose • Monosaccharide analysis: Glucose/mannose/galacturonic acid/ Not tested Emulsifying activity Caruso et al. (2017) pH 7.0) arabinose/galactose/glucosamine/glucuronic acid (1/0.5/0.3/ Cryoprotective activity 0.25/0.1/0.1/0.1, by relative proportion) Heavy metal chelation Winogradskyella sp. CAL396 (4 °C, Sucrose • Monosaccharide analysis: Mannose/arabinose/galacturonic Not tested Cryoprotective activity Caruso et al. (2017) pH 7.0) acid/glucuronic acid/galactose/glucose/glucosamine (1/0.9/ Heavy metal chelation 0.4/0.3/0.2/0.2/0.01, by relative proportion) Colwellia sp. GW185 (15 °C, pH Sucrose • Monosaccharide analysis: Glucose/mannose/galactose/ Not tested Cryoprotective activity Caruso et al. (2017) 6.0) galactosamine/glucuronic acid/galacturonic acid (1/1/0.7/0.7/ Heavy metal chelation 0.3/0.04, by relative proportion) Shewanella sp. CAL606 (4 °C, pH Sucrose • Monosaccharide analysis: Glucose/galactose/mannose/ Not tested Cryoprotective activity Caruso et al. (2017) 7.0) galactosamine/glucuronic acid/galacturonic acid (1/1/0.9/0.6/ Heavy metal chelation 0.3/0.1, by relative proportion) Colwellia psychrerythraea 34H Not added • The repeating unit: structure with a N-acetyl- Not tested Inhibitory effect on ice recrystallization Casillo, Parrilli et al. (2017), (4 °C, pH 7.6) quinovosamine and two galacturonic acid residues Cryoprotective activity for the strain itself Marx et al. (2009) Pseudoalteromo-nas ulvae TC14 Not added EPS 1 EPS 1 Anti-biofilm activity Brian-Jaisson et al. (2016) (20 °C, pH 7.6) • Monosaccharide analysis: Glucose 1000 kDa EPS 2 EPS 2 • Monosaccharide analysis: Glucose 4000 kDa

11 Pseudoalteromo-nas elyakovii Arcpo Glucose • Monosaccharide analysis: Mannose/galacturonic acid (3.3/1.0, 17,000 kDa Cryoprotective activity Kim, Kim, Park, and Yim, 2016) 15 (15 °C, pH 7.2) by relative molar ratio) Pseudomonas sp. ID1 (11 °C, pH Glucose • Monosaccharide analysis: Glucose/galactose/fucose (50.38/ Above 2000 Emulsifying activity Carrión et al. (2015) 7.0) 25.34/24.28, by weight percentage) kDa Cryoprotective activity for the strain itself as well as for other bacteria Pseudoplastic rheological property Cobetia marina DSMZ 4741 (20 °C, Glucose • Monosaccharide analysis: Ribose/3-deoxy-D-manno-oct-2- 270 kDa Not tested Lelchat et al. (2015) pH 7.6) ulosonic acid (1/1, by molar ratio) Polaribacter sp. SM1127 (15 °C, pH Glucose • Monosaccharide analysis: N-acetylglucosamine/mannose/ 220 kDa Antioxidant activity Sun et al. (2015) 7.0) Glucuronic acid/galactose/fucose/glucose/rhamnose (28.0/ Moisture-retention ability 23.4/21.4/17.3/7.4/1.6/0.8, by molar percentage) Pseudoplastic rheological property Low-temperature protective effect on human dermal fibroblasts Nontoxic and nonirritating to skin Pseudoalteromo-nas sp. SM20310 Glucose • Monosaccharide analysis: Mannose/glucose/galactose/N- Above 2000 Enhance the high-salinity tolerance for the strain itself Liu et al. (2013) (15 °C, pH 7.5) acetylglucosamine/rhamnose/N-acetylgalactosamine/xylose kDa Cryoprotection for the strain itself and other bacteria (71.7/10.7/9.0/4.0/2.1/1.5/0.9, by molar percentage) Pseudoalteromo-nas sp. S-5 (8 °C, Glucose • Monosaccharide analysis: Glucose/galactose/mannose (50.9/ 397 kDa Immunoregulatory activity Bai et al. (2012) pH 7.6) 44.3/4.8, by molar ratio) Carbohydrate Polymers205(2019)8–26 Pseudoalteromo-nas sp. SM9913 Lactose • Monosaccharide analysis: 6-Glucose, terminal arabinofuranosyl, 40 kDa Function stabilizing and thermostability enhancement Qin, Zhu, Chen, Wang, and Zhang, (15 °C, pH 7.5) terminal glucopyranosyl, terminal galactose, 4-xylose, 4-glucose on the proteases secreted by the same strain 2007) and 3,6-galactose (61.8/11.0/11.2/3.1/3.9/5.0/4.0, by weight Metal-binding property percentage) Flocculation property • The linkage between the repeating sugar units: α-1,6 linkage, and this EPS was structurally characterized as a linear arrangement of α-(1,6)-glucose and a high degree of acetylation • The repeating unit: -6)-[3,6-O-acetyl]-α-D-Glcp-(1-6)-[3-O- acetyl]-α-D-Glcp-(1-6)-[3-O-acetyl]-α-D-Glcp-(1-6)-[3-O- acetyl]-α-D-Glcp-(1- (continued on next page) .Wn tal. et Wang J. Table 1 (continued)

Extremophiles Sugar carbon Monosaccharide composition and linkage pattern Molecular Properties and activities Reference (specific conditions) source weight

Flavobacterium frigidarium Glucose • Monosaccharide analysis: Arabinose/mannose/galactose/ 1810 kDa Cryoprotectant for microorganisms Nichols, Lardière et al. (2005) CAM005 (20 °C, pH 7.0) glucose/glucuronic acid/N-acetyl-glucosamine (5/74/3/8/8/1, w/w at percentage of total sugars) Myroides odoratus CAM030 (20 °C, Glucose • Monosaccharide analysis: Arabinose/rhamnose/xylose/ 190 kDa Cryoprotectant for microorganisms Nichols, Lardière et al. (2005) pH 7.0) mannose/galactose/glucose/galacturonic acid/glucuronic acid/ N-acetylgalactosamine/N-acetylglucosamine (6/1/2/48/4/9/2/ 10/10/8, w/w at percentage of total sugars) Polaribacter irgensii CAM006 Glucose • Monosaccharide analysis: Arabinose/fucose/mannose/ 2100 kDa Cryoprotectant for microorganisms Nichols, Lardière et al. (2005) (20 °C, pH 7.0) galactose/glucose/glucuronic acid/N-acetylgalactosamine/N- acetyl-glucosamine (2/11/33/38/4/6/1/4, w/w at percentage of total sugars) Pseudoalteromo-nas sp. CAM003 Glucose • Monosaccharide analysis: Arabinose/ribose/rhamnose/fucose/ 1800 kDa Cryoprotectant for microorganisms Nichols, Lardière et al. (2005) (20 °C, pH 7.0) mannose/glucose/glucuronic acid/N-acetylgalactosamine/N- acetylglucosamine (4/2/6/29/40/16/1/1/1, w/w at percentage of total sugars) Pseudoalteromo-nas sp. CAM015 Glucose • Monosaccharide analysis: Arabinose/rhamnose/xylose/ 2800 kDa Cryoprotectant for microorganisms Nichols, Lardière et al. (2005) (20 °C, pH 7.0) mannose/galactose/glucose/glucuronic acid/N-acetyl- galactosamine (10/6/1/36/4/38/3/3, w/w at percentage of total sugars) Pseudoalteromo-nas sp. CAM023 Glucose • Monosaccharide analysis: Arabinose/mannose/galactose/ 1800 kDa Cryoprotectant for microorganisms Nichols, Lardière et al. (2005) (20 °C, pH 7.0) glucose/galacturonic acid/glucuronic acid/N-acetyl- galactosamine/N-acetyl-galactosamine (12/2/1/75/5/3/2, w/w at percentage of total sugars) Pseudoalteromo-nas sp. CAM025 Glucose • Monosaccharide analysis: Arabinose/ribose/rhamnose/fucose/ 5700 kDa Cryoprotectant for microorganisms Nichols, Lardière et al. (2005)

12 (20 °C, pH 7.0) mannose/galactose/glucose/galacturonic acid/N-acetyl- galactosamine (3/1/5/1/1/5/52/30/1, w/w at percentage of total sugars) Pseudoalteromo-nas sp. CAM036 Glucose • Monosaccharide analysis: Arabinose/mannose/galactose/ 1700 kDa Cryoprotectant for microorganisms Nichols, Lardière et al. (2005) (20 °C, pH 7.0) glucose/galacturonic acid/N-acetyl-galactosamine/N-acetyl- glucosamine (3/24/1/26/30/14/1, w/w at percentage of total sugars) Pseudoalteromo-nas sp. CAM064 Glucose • Monosaccharide analysis: Arabinose/mannose/galactose/ 100 kDa Cryoprotectant for microorganisms Nichols, Lardière et al. (2005) (20 °C, pH 7.0) glucose/glucuronic acid/N-acetyl-galactosamine/N-acetyl- glucosamine (4/64/4/8/6/11/2, w/w at percentage of total sugars) Shewanella livingstonensis CAM090 Glucose • Monosaccharide analysis: Arabinose/rhamnose/xylose/ 80 kDa Cryoprotectant for microorganisms Nichols, Lardière et al. (2005) (20 °C, pH 7.0) mannose/galactose/glucose/glucuronic acid/N- acetylgalactosamine (13/2/2/41/5/10/20/7, w/w at percentage of total sugars) Pseudoalteromo-nas haloplanktis Not added • Monosaccharide analysis: Mannose/glucose (1/trace, by relative Not tested Not tested Corsaro et al. (2004) TAC 125 (15 °C, pH 7.5) ratio) Pseudomonas sp. NCMB 2021 Glucose EPS 1 Not tested Metal cation precipitation Christensen, Kjosbakken, and Smidsrød, (17 °C, pH 7.5) • Monosaccharide analysis: Glucose/galactose/glucuronic acid/ 1985) Carbohydrate Polymers205(2019)8–26 galacturonic acid (1/0.81/0.42/0.32, by molar ratio) EPS 2 • Monosaccharide analysis: N-acetylglucosamine/2-keto-3- deoxyoctulosonic acid/unidentified 6-deoxyhexose (1/1/1, by molar ratio) Halothermophiles

(continued on next page) .Wn tal. et Wang J. Table 1 (continued)

Extremophiles Sugar carbon Monosaccharide composition and linkage pattern Molecular Properties and activities Reference (specific conditions) source weight

Halomonas nitroreducens WB1 Glucose EPS 1 EPS 1 5200 Emulsifying activity Chikkanna, Ghosh, and Kishore, (2018) (60 °C, pH 7.5, 5% w/v NaCl) • Monosaccharide analysis: Glucose/mannose/galactose/xylose kDa Antioxidant activity (28/64/6/traces, by weight percentage) EPS 2 Heavy metal binding capacity EPS 2 30 kDa Pseudoplastic rheological property • Monosaccharide analysis: Glucose/mannose/rhamnose/ EPS 3 arabinose/xylose (18.5/44/2/1.5/traces, by weight percentage) 1.3 kDa EPS 3 • Monosaccharide analysis: Glucose/mannose/galactose/ galacturonic acid/fructose (19/56.5/14.2/1.5/traces, by weight percentage) Bacillus licheniformis B3-15 (45 °C, Glucose EPS 1 EPS 2 Antiviral and immunoregulatory activities Spanò and Arena (2016), Arena et al. pH 7.0, 2% w/v NaCl) • Monosaccharide analysis: Mannose/glucose (1.0/0.3, by molar 600 kDa (2006), ratio) Maugeri et al. (2002) EPS 2 • Monosaccharide analysis: Mannose • Repeating unit: Tetrasaccharide EPS 3 • Monosaccharide analysis: Glucose Bacillus licheniformis T14 (50 °C, Sucrose • Monosaccharide analysis: Fructose/fucose/glucose/ 1000 kDa Degradation temperature Spanò, Laganà, Visalli, Maugeri, and pH 8.0, 5% w/v NaCl) galactosamine/mannose (1.0/0.75/0.28/trace/trace, by relative 240 °C Gugliandolo, (2016), molar ratio) Anti-cytotoxicity Gugliandolo et al. (2013), • Saccharide repeating unit: Trisaccharide unit Viscoelasticity Spanò et al. (2013) • Anomeric configuration: β-manno-pyranosidic configuration Antiviral and immunomodulatory effects against herpes simplex virus type 2 (HSV-2)

13 Anti-biofilm activity Emulsifying and stabilizing activities Geobacillus sp. 1A60 (50 °C, pH Sucrose • Monosaccharide analysis: Mannose/galactose/galactosamine/ Not tested Heavy metal binding capacity Gugliandolo, Lentini, Spanò, and 8.0, 5% w/v NaCl) fucose/glucose (1/0.69/0.65/0.59/0.35, by relative proportion) Maugeri, 2012) Halophiles Chromohalobac-ter canadensis 28 Lactose • Monosaccharide analysis: Glucosamine/glucose/rhamnose/ Above 1000 Degradation temperature Radchenkova et al. (2018) (30 °C, pH 8.5, 15% w/v xylose/unknown sugar (36.7/32.3/25.4/1.7/3.9, by weight kDa 250 °C NaCl) percentage) Pseudoplastic rheological property High swelling behavior Emulsifying and stabilizing activities Foaming ability Halolactibaci-llus miurensis SEEN Glucose • Monosaccharide analysis: Galactose/glucose/xylose/fructose/ Not tested Antioxidant activity Arun et al. (2017) MKU3 (32 °C, pH 8.0, 75 g/L mannose/rhamnose (61.87/25.17/not tested/not tested/not NaCl) tested/not tested, by relative percentage) Kocuria rosea ZJUQH (30 °C, pH Not added • Monosaccharide analysis: Glucose 56.59 kDa Not tested Gu, Jiao, Wu, Liu, and Chen, 2017) 7.0, 5.8% w/v MgSO4) Vibrio alginolyticus CNCM I-4994 Glucose • Monosaccharide analysis: Galacturonic acid/N-acetyl- 1160 kDa Not tested Drouillard et al. (2015)

(25 °C, pH 7.2, 30 g/L sea glucosamine (3/1, by relative ratio) Carbohydrate Polymers205(2019)8–26 salts) Halomonas smyrnensis AAD6T Sucrose • Monosaccharide analysis: Fructose Above 1000 Degradation temperature 253 °C Sarilmiser and Oner (2014), (37 °C, pH 7.0, 137.2 g/L kDa Bioflocculating activity Küçükaşik et al. (2011), NaCl) Anti-cytotoxicity Sam et al. (2011), Biocompatibility Poli et al. (2009) Antitumor activity after periodate oxidation Alteromonas macleodii (28 °C, pH Glucose • Monosaccharide analysis: Galactose/glucose/rhamnose/ 1100 kDa Not tested Le Costaouëc et al. (2012) 7.2, 30 g/L sea salts) glucuronic acid/galacturonic acid/mannose/fucose (5.9/2.6/ 2.5/2.0/1.9/1.4/1.0, by molar ratio) (continued on next page) .Wn tal. et Wang J. Table 1 (continued)

Extremophiles Sugar carbon Monosaccharide composition and linkage pattern Molecular Properties and activities Reference (specific conditions) source weight

Halomonas almeriensis M8T (32 °C, Glucose EPS 1 EPS 1 Emulsifying activity Llamas et al. (2012) pH 7.0, 7.5% w/v total salts) • Monosaccharide analysis: Mannose/glucose/rhamnose (72/ 6300 kDa Heavy metal binding capacity 27.5/0.5, by weight percentage) EPS 2 Pseudoplastic rheological property EPS 2 15 kDa • Monosaccharide analysis: Mannose/glucose (70/30, by weight percentage) Vibrio sp. QY101 (25 °C, pH 7.0, Alginate • Monosaccharide analysis: Rhamnose/galacturonic acid/ 546 kDa Biofilm formation inhibition activity Jiang et al. (2011) 3.0% w/v NaCl) glucuronic acid/glucosamine/galactose/glucose/fucose/ Pre-existing biofilm disruption activity mannose (23.90/23.05/21.47/12.15/6.89/6.57/3.61/2.36, by molar percentage) Halomonas stenophila B100 (32 °C, Glucose • Monosaccharide analysis: Glucose/galactose/mannose (44.5/ 375 kDa Antitumor activity after oversulphation Ruiz-Ruiz et al. (2011) pH 7.2, 7.5% w/v marine 40.5/15.0, by weight percentage) salts) Halomonas stenophila N12T (32 °C, Glucose • Monosaccharide analysis: Glucose/fucose/mannose (48.82/ 250 kDa Antitumor activity after oversulphation Ruiz-Ruiz et al. (2011) pH 7.2, 7.5% w/v marine 25.69/25.47, by weight percentage) salts) Salipiger mucosus A3T (32 °C, pH Glucose • Monosaccharide analysis: Mannose/galactose/glucose/fucose 250 kDa Emulsifying activity Llamas et al. (2010) 7.0, 2.5% w/v total salts) (34/32.9/19.7/13.4, by weight percentage) Heavy metal binding capacity Pseudoplastic rheological property Idiomarina fontislapidosi F23T Glucose EPS 1 EPS 1 1500 Emulsifying activity Mata et al. (2008) (32 °C, pH 7.2, 7.5% w/v total • Monosaccharide analysis: Mannose/glucose/galactose/xylose kDa Heavy metal binding capacity salts) (46.35/28.25/14.85/trace, by molar percentage) EPS 2 Pseudoplastic rheological property EPS 2 15 kDa • Monosaccharide analysis: Mannose/glucose/galactose/xylose

14 (40/40/20/trace, by molar percentage) Idiomarina ramblicola R22T (32 °C, Glucose EPS 1 EPS 1 Emulsifying activity Mata et al. (2008) pH 7.2, 7.5% w/v total salts) • Monosaccharide analysis: Mannose/glucose/rhamnose (68.2/ 550 kDa Heavy metal binding capacity 25/6.8, by molar percentage) EPS 2 Pseudoplastic rheological property EPS 2 20 kDa • Monosaccharide analysis: Mannose/galacturonic acid/ glucose/ rhamnose/xylose (53.6/25.29/18.9/trace/trace, by molar percentage) Alteromonas hispanica F32T (32 °C, Glucose • Monosaccharide analysis: Mannose/glucose/xylose/rhamnose 19,000 kDa Emulsifying activity Mata et al. (2008) pH 7.2, 7.5% w/v total salts) (62.75/18.15/12.25/6.85, by molar percentage) Heavy metal binding capacity Pseudoplastic rheological property Halomonas eurihalina F2-7 (32 °C, Glucose • Monosaccharide analysis: Glucose/mannose/rhamnose (2.9/ Not tested Emulsifying activity Martínez-Checa, Toledo, El Mabrouki, pH 7.2, 7.5% w/v total salts) 1.5/1, by relative ratio) Pseudoplastic rheological property Quesada, and Calvo, (2007), Bejar, Calvo, Moliz, Diaz-Martinez, and Quesada, (1996) Halomonas ventosae A112T (32 °C, Glucose • Monosaccharide analysis: Glucose/mannose/galactose (1.75/4/ 53 kDa Emulsifying activity Mata et al. (2006) pH 7.2, 7.5% w/v total salts) 1, by molar ratio), and small quantities of xylose, arabinose and Heavy metal binding capacity galacturonic acid Biofilm formation capacity Pseudoplastic rheological property Carbohydrate Polymers205(2019)8–26 Halomonas ventosae A116 Glucose • Monosaccharide analysis: Glucose/mannose/galactose (1.25/4/ 52 kDa Emulsifying activity Mata et al. (2006) (32 °C, pH 7.2, 7.5% w/v total 1, by molar ratio), and small quantities of xylose, arabinose and Heavy metal binding capacity salts) galacturonic acid Biofilm formation capacity Pseudoplastic rheological property Halomonas anticariensis Glucose • Monosaccharide analysis: Glucose/mannose/galacturonic acid 20 kDa Emulsifying activity Mata et al. (2006) FP35T (32 °C, pH 7.2, 7.5% w/ (1/3/2.5, by molar ratio) Heavy metal binding capacity v total salts) Biofilm formation capacity Pseudoplastic rheological property (continued on next page) .Wn tal. et Wang J. Table 1 (continued)

Extremophiles Sugar carbon Monosaccharide composition and linkage pattern Molecular Properties and activities Reference (specific conditions) source weight

Halomonas anticariensis FP36 Glucose • Monosaccharide analysis: Glucose/mannose/galacturonic acid 46 kDa Emulsifying activity Mata et al. (2006) (32 °C, pH 7.2, 7.5% w/v total (1/2.5/2.2, by molar ratio) Heavy metal binding capacity salts) Biofilm formation capacity Pseudoplastic rheological property Halomonas maura S-30 (32 °C, pH Glucose • Monosaccharide analysis: Mannose/galactose/glucose/ 4700 kDa Heavy-metal uptake Arias et al. (2003) 7.0, 2.5% w/v sea salts) glucuronic acid (34.8/14/29.3/21.9, by weight percentage) Viscosifying potential Pseudoplastic rheological property Aphanothece halophytica GR02 Not added EPS 1 EPS 2 Gelling property Li et al. (2001) (30 °C, pH 7.0, 1 M NaCl) • Monosaccharide analysis: Arabinose/rhamnose/fucose/ Above 2000 Strong affinity for metal ions mannose/glucose/galactose (trace/0.06/0.05/0.08/1/0.75, by kDa molar ratio) and glucuronic acid 3.58% of dry weight EPS 2 • Monosaccharide analysis: Arabinose/fucose/mannose/glucose (1/2.08/1.57/2.87, by molar ratio) and glucuronic acid 15.78% of polysaccharide dry weight Alteromonas sp. 1644 (25 °C, pH Fructose • Monosaccharide analysis: Galactose/glucose/glucuronic acid/3- Not tested Gelling property Samain, Miles, Bozzi, Dubreucq, and 7.0, 30 g/L sea salts) O-[(R)-1-carboxyethyl]-D-glucuronic acid/galacturonic acid Rinaudo, 1997) (1.0/0.92/0.7/0.34/0.26, by molar ratio) Haloalkaliphiles

Halomonas sp. CRSS (30 °C, pH Acetatea • Monosaccharide analysis: Glucose/fructose/glucosamine/ Not tested Viscosity above 0.5 η Poli et al. (2004) 9.0, 100 g/L NaCl) galactosamine (1/0.7/0.3/trace, by relative proportion) Solution viscosity can increase at pH 2-3 with 2.5% (w/v) NaCl Bacillus sp. (37 °C, pH 10.5, 40 g/L Glucose • Monosaccharide analysis: D-galactopyranuronic acid, 2,4- Not tested Not tested Corsaro, Grant, Grant, Marciano, and 15 NaCl) diacetamido-2,4,6-trideoxy-D-glucopyranose, 2-acetamido-2- Parrilli, (1999), Duckworth, Grant, deoxy D-mannopyranuronic acid and D-galactopyranuronic acid Jones, and Van Steenbergen, 1996) with the carboxyl group amide-linked to glycine • The repeating unit: -3)-a-d-GalpA(Gly)-(1-4)-b-d-ManpNAcA-(1- 4)-a-d-GalpA-(1-3)-a-d-QuipNAc4NAc-(1- Alkaliphiles Cronobacter sakazakii (30 °C, pH Sucrose • Monosaccharide analysis: Glucose/mannose/galactose/xylose/ 3760 kDa Degradation temperature 280 °C Jain et al. (2012) 10) arabinose (14/24/14/20/1.9, by weight percentage) Pseudoplastic rheological property Emulsifying activity Bacillus cereus (23 °C, pH 10.5) Glucose • Monosaccharide analysis: Arabinose/xylose/mannose/ Above 167 Calcite binding Perry et al. (2005) galactose/glucose/N-acetylglucosamine (5.0/3.4/70.3/12.1/ kDa 4.7/4.5, by molar percentage) Bacillus thuringiensis (23 °C, pH Glucose • Monosaccharide analysis: Arabinose/rhamnose/xylose/ Above 167 Calcite binding Perry et al. (2005) 10.5) galacturonic acid/mannose/ galactose/glucose (9.4/3.2/5.6/ kDa 7.5/52.2/16.9/5.2, by molar percentage)

a Non-sugar carbon source. Carbohydrate Polymers205(2019)8–26 J. Wang et al. Carbohydrate Polymers 205 (2019) 8–26 microorganisms in a marine environment may provide both cryopro- capacity, and rheological properties, with existing and potential ap- tection and a buffering effect against low temperature and high salinity plications in a range of industrial fields, such as utilization as a sub- simultaneously. The secondary molecular structure analysis of psy- stitute for xanthan gum in the food industry. chrophilic EPS indicates that a pseudohelicoidal structure may be ad- Changes in salinity affects the biosynthesis of halophilic EPSs, vantageous for the inhibition of ice recrystallization (Casillo, Parrilli especially the ratio for each type of monosaccharide composition. To et al., 2017). Moreover, the decoration by amino acid motifs onto the protect the microorganism from increasing salinity, the content of some monosaccharide moieties was speculated to endow a structural equili- monosaccharide components in EPS may need to be modified in order brium between hydrophilic and hydrophobic regions in the EPS mole- to maintain its functions. For the EPS obtained from strain Aphanothece cule, and thus contribute to the inhibitory effect on ice crystal devel- halophytica GR02, the proportions of galactose and rhamnose decreased opment (Casillo, Ståhle et al., 2017). The sulphate moieties in when the NaCl concentration in the medium was elevated from 0.5 to psychrophilic EPSs may also play a significant role against extremely 2.0 M; in contrast, the proportions of arabinose and glucose increased cold environments (Nichols, Guezennec et al., 2005; Nichols, Bowman with NaCl concentration. Meanwhile, the present in et al., 2005). The physical, rheological, and chemical properties of EPSs the EPS at different salinities stayed the same (P. Li, Liu, & Xu, 2001). can be influenced by the length of the polymer chain, and the high This indicates that the increase of glucose and arabinose, and the de- molecular weights of EPSs from psychrophiles provide greater oppor- crease of galactose and rhamnose in the EPS secreted by Aphanothece tunity for complex entanglement of polymer chains and intramolecular halophytica GR02 may be advantageous to its survival in a high salinity associations, which may contribute to the tertiary structure and en- environment. Mata et al. (2006) mentioned that for the strain Halo- hance the physical behavior of the EPSs in their environment (Nichols, monas ventosae A112T, its EPS incorporated a significant quantity of Bowman et al., 2005). Besides, the EPSs with higher molecular weight sulphate. Sulphate is not commonly found in mesophilic EPSs; however, also possess better water binding capacity than EPSs with lower mo- it has been observed in the EPSs excreted by microorganisms living in lecular weight (Prechtl, Wefers, Jakob, & Vogel, 2018). saline habitats. In addition, the EPSs from halophiles usually contain Normally, psychrophilic EPS production can be inhibited by a re- significant amounts of uronic acids. The high viscosity of the EPS so- latively elevated temperature, in the region of 20 °C and above (Nichols, lution at acidic pH and the gelification capacity may be due to the high Garon, Bowman, Raguenes, & Guezennec, 2004, 2005). The contents of uronic acid content (Béjar, Llamas, Calvo, & Quesada, 1998). EPSs with monosaccharide components in psychrophilic EPSs can be modified high concentrations of charged components (e.g. uronic acids) often through change of temperature, and some of the monosaccharides and form gels in the presence of metal ions and have enormous potential for other residues in EPSs from psychrophiles may help to confer ad- removing toxic metal from polluted environments and wastewater as an vantageous cryoprotectant properties. For example, the uronic acid alternative to other physical and chemical methods. content in the EPSs produced by Pseudoalteromonas sp. CAM025 at −2 °C and 10 °C was significantly higher than that at 20 °C; and the 2.4. EPSs produced by acidophiles monosaccharide compositions were also found to differ among the EPSs harvested at −2 °C, 10 °C, and 20 °C (Nichols, Bowman et al., 2005). Acidophiles are extremophiles which inhabit a low pH environment, The psychrotolerant strain Lactobacillus sakei TMW 1.411 produced usually less than pH 3 for optimum growth. Some of the acidophiles dextran with less branching and higher molecular weight at 10 °C than cannot grow at all in a neutral pH condition (Baker-Austin & Dopson, the dextran produced at 30 °C (Prechtl et al., 2018). At temperatures 2007; Johnson, 1998; Johnson, Joulian, d’Hugues, & Hallberg, 2008). below the optimum temperature for cell growth, the psychrophiles were Both natural and artificial acidic niches can occur in the biosphere, such stimulated to produce excessive EPSs (Marx, Carpenter, & Deming, as a sulfidic mine area or a marine volcanic vent. The acidic environ- 2009; Nevot et al., 2008; Nichols et al., 2004). This is consistent with ments usually include the presence of sulphur, sulphide, and their the fact that EPS production is one of the main mechanisms to protect oxidates. Pyrite is one of the main acidic niches for acidophiles. These extremophiles and enable them to survive in extreme conditions. areas are quite toxic due to high concentrations of various heavy metal Therefore, output of EPS for each cell can be enhanced with the dete- sulphides, but they are rich in valuable metals, such as Fe, Cu, Co, Al, rioration of environmental conditions in a certain range, albeit the cell Mg, Zn, and Mn (Dopson, Baker-Austin, Koppineedi, & Bond, 2003; Jiao growth may sharply decrease. Enhancing net EPS production may thus et al., 2010; Johnson et al., 2008; Nicolaus et al., 2010). involve identifying the optimal trade-off between increased EPS pro- Compared with the research for other kinds of extremophilic EPSs, duction per cell and reduced cell count. the acidophilic EPSs have not been studied sufficiently to reveal their In several former studies, the stabilization effect of psychrophilic fermentation process, molecular structure, or properties. Usually EPSs EPS for protease against thermal denaturation was confirmed (Huston, from acidophiles are considered as bioproducts generated in another Methe, & Deming, 2004; Junge, Eicken, Swanson, & Deming, 2006; bioprocessing technology such as a bioleaching process. For acid- Marx et al., 2009), which indicates that psychrophilic biopolymers can ophiles, the genome analysis cannot identify ubiquitous DNA adapta- be applied to the stabilization of industrially promising enzymes used in tions for growth in an extremely low pH environment (Baker-Austin & unfavorable conditions. In future research on psychrophilic EPSs, it is Dopson, 2007). On the other hand, the EPSs produced by acidophiles recommended that significant insights may be found by comparing the may play a protective role against stress conditions related to the low structure and function of EPSs from different culture conditions, in pH and presence of metals. Acidophilic EPS biosynthesis can be in- order to reveal what kind of structure can be more advantageous for hibited by increased temperature during the bioleaching process, and protection and stabilization effects. the inhibited EPS production may have been related to the loss of bioleaching efficiency observed in the reactor when the temperature 2.3. EPSs produced by halophiles was increased (d’Hugues et al., 2008). This phenomenon indicates that the acidophilic EPSs protecting acidophiles from an acidic environment Moderately halophilic bacteria are defined as those which grow are not able to protect them against a relatively high temperature optimally in media containing 5–20% (w/v) salts, and they constitute condition, unlike thermophilic EPSs. Therefore, it is of significant in- the most important eubacteria group living in hypersaline habitats terest to explore the acidophilic EPSs for functional diversity elucida- (Ollivier, Caumette, Garcia, & Mah, 1994;C.Qian et al., 2018). Most tion through molecular level structure and comparison among acid- halophilic EPSs are heteropolysaccharides, and mannose and glucose ophilic and other extremophilic EPSs as models. are the most common monosaccharide moieties in halophilic EPSs Some acidophilic EPSs were discovered during the study of extra- (Table 1). So far, the research focus for halophilic EPSs properties has cellular polymeric substances, which are one of the major components been emulsifying activity, gelling properties, heavy metal binding in biofilms, and they mainly consist of EPSs, proteins, and nucleic acids

16 J. Wang et al. Carbohydrate Polymers 205 (2019) 8–26

(Flemming & Wingender, 2010; Moreno-Paz, Gómez, Arcas, & Parro, costs, allowing extremophilic EPSs to compete in the biopolymer 2010; Subramanian, Yan, Tyagi, & Surampalli, 2010; Vu, Chen, market. Several biopolymers from mesophiles and neutrophiles, such as Crawford, & Ivanova, 2009). The extracellular polymeric substances , alginate, gellan, and sphingan have already been profoundly containing acidophilic EPSs are usually generated by mixed cultures studied for the enzymes and genes involved in their biosynthetic during the bioleaching process. Bioleaching uses the oxidation ability of pathways (Ates, 2015; Schmid, Sieber, & Rehm, 2015). However, the bacteria to dissolve metal sulphides in order to facilitate the extraction information about metabolic pathways and functional assignment of and recovery of precious metals from primary ores and concentrates. gene clusters for extremophilic EPS biosynthesis is still limited. Re- The involved microbial consortia are mainly composed of acidophilic, search focusing on genetically modified strains capable of producing autotrophic iron-oxidizing, and sulphur-oxidizing bacteria (Michel highly improved levels of extremophilic EPS is also necessary since, et al., 2009). In Zeng’s report (Zeng et al., 2010), an acidophilic mixed compared to mesophilic and neutrophilic EPS-producing strains, ex- culture was able to produce extracellular polymeric substances during tremophilic bacteria are relatively inefficient at producing EPSs. the bioleaching process, and Acidithiobacillus caldus and Leptospirillum ferriphilum were considered as the dominant microorganisms in the 3.1. Engineering strategies in EPS biosynthetic pathways for improved EPS mixed culture. The extracellular polymeric substance had protein, production and modified molecular weight polysaccharide, fatty acid, and ferric ion as its main components. Rhamnose, fucose, xylose, mannose, glucose, and uronic acids were the EPS biosynthesis is highly associated with catabolic processes of components of the polysaccharide which could be considered to come oxidation and does not interfere with other anabolic bioprocesses from the EPS excreted by the mixed culture during the bioleaching (Chawla et al., 2009). As a carbon and energy intensive process, the process. The percentages of these components varied at different sam- biosynthesis of extremophilic EPSs usually requires the recruitment of pling time during bioleaching, while the presence of these components nucleoside diphosphate saccharides (NDP-sugars) as precursors, glyco- remained stable. A pure culture, Thiobacillus ferrooxidans, was also syltransferases (GTs) for assembly, and membrane proteins for the carried in the bioleaching process, and the monosaccharide units of the transfer of repeat units across cell envelope. Generally, the EPS bio- carbohydrate in the extracellular polymeric substance were rhamnose, synthetic pathway starts from glycolysis of simple sugar for cytosolic fucose, xylose, mannose, glucose, and glucuronic acid. This composi- formation of the NDP-sugar precursors; then the monosaccharides are tion varied greatly when T. ferrooxidans was grown in a different sequentially transported from nucleotide-sugar donors to activated lipid medium containing iron (II) sulphate, pyrite, or sulfur as the solid carriers and assembled as repeating units of polysaccharide through substrate (Gehrke, Telegdi, Thierry, & Sand, 1998). GTs. Finally, the EPS needs to be exported to an extracellular en- vironment. Based on the general biosynthetic pathway of EPS, the genes 2.5. EPSs produced by alkaliphiles involved can be organized into three functional types: (1) genes in- volved in NDP-sugar synthesis, (2) genes coding for GTs required for The alkaliphiles are microorganisms that grow optimally or very biosynthesis of EPS repeating unit, (3) genes encoding proteins for well at pH values above 9, often between 10 and 12, but cannot grow or polymerization and export (Ates, 2015). grow slowly at near-neutral pH values (Horikoshi, 1999). Soda lakes During the first phase of EPS biosynthesis, the NDP-sugars represent and deserts represent the most stable, naturally occurring alkaline en- the interface between primary and secondary metabolism (Ates, Arga, vironments which can be found all over the world (Rees, Grant, Jones, & Oner, 2013). A bottleneck is the low level of activated NDP-sugar & Heaphy, 2004). The enzymes isolated from alkaliphiles, including precursors which can be exploited as design space through metabolic alkaline proteases, amylases, cellulases, and lipase, have been applied engineering to alter the expression of enzymes involved in the central in various industrial sectors such as the detergent industry (Ito et al., metabolism for supplying nucleotide-sugar precursors. The higher EPS 1998). As with other kinds of extremophiles, the alkaliphiles produce producing mutant demonstrated that the specific activities of phos- EPSs as metabolic products. So far, certain functions of EPSs from al- phoglucomutase, UDP-glucose pyrophosphorylase, UDP-glucose dehy- kaliphiles have been partially studied (Table 1), but more research on drogenase, and UDP-galactose-4-epimerase were higher than those in molecular structure, properties, and the biosynthesis pathway of alka- the wild-type strain (Fig. 2), indicating these enzymes involved in NDP- liphilic EPSs are necessary to improve scientific understanding and to sugar synthesis can be potential targets for enhancement of EPS pro- enable targeted industrial applications. duction (Li et al., 2010; Welman, Maddox, & Archer, 2006; Zhu et al., Alkaliphilic EPSs are functional for the attachment of the associated 2014). Although it is still relatively nascent for extremophilic bacteria microbial strains to a certain matrix. For example, the binding strength to be applied as intact platforms for metabolic engineering, a group of to calcite was found to be due to the chemical properties of the EPSs extremophiles have already been metabolically engineered for en- secreted by two natural alkaliphiles isolated from biofilms on historic hanced biofuel or enzyme production due to the recent expansion of the limestone. Meanwhile, these two alkaliphilic EPSs could also contribute genetic systems and tools for extremophiles (Lin & Xu, 2013; Zeldes to calcite dissolution in the biofilm development process (Perry et al., et al., 2015). In EPS-producing bacteria, the sugar substrates are either 2005). Unlike most other extremophiles, for which sugar is the optimal converted into EPS synthesis or cell mass by alternative intermediary carbon source for EPS production, the most efficient carbon source for metabolic routes. The rerouting of the carbon flux through the aug- EPS production of the haloalkalophilic strain Halomonas sp. CRSS was mentation of a critical enzyme at the principal branch point to NDP- acetate. The growth conditions strongly influenced the cumulative sugar synthesis was considered as a strategy to enhance the EPS pro- production, relative fractions of different monosaccharides, and duction of several mesophilic bacteria. The homologous over-expres- monomer compositions of the EPS from Halomonas sp. CRSS (Poli et al., sion of phosphoglucomutase in Sphingomonas sanxanigenens strain re- 2004). sulted in a 17% increase in EPS production (Huang et al., 2013). However, the flow of carbon towards the synthesis of EPS by Sphingo- 3. Metabolic and genomic engineering of extremophilic EPSs monas sp. strain S7 was manipulated by augmenting the cellular phosphoglucomutase activity with additional genes, and no significant Extremophilic EPSs have increasing significance in material and increase in EPS yield was observed (Thorne, Mikolajczak, Armentrout, biomedical applications that require a more profound understanding of & Pollock, 2000). The over-expression of UDP-glucose pyropho- the metabolic pathways and biosynthetic mechanisms of EPS in order to sphorylase involved in the synthesis of UDP-glucose also had negligible control the production process and molecular structure, and hence the effect on EPS productivity. Meanwhile, the inactivation of glucose-6- physiochemical properties. The development of engineered EPS-pro- phosphate dehydrogenase could not divert carbon flow toward EPS ducing strains can also reduce their exceptionally expensive production synthesis (Sá-Correia et al., 2002). On the other hand, the simultaneous

17 J. Wang et al. Carbohydrate Polymers 205 (2019) 8–26

Fig. 2. Potential engineering targets for increasing monosaccharide precursors for EPS biosynthesis in extremophiles. PTS: phosphotransferase system; ABC: ATP- binding cassette transporter; GDP: guanosine diphosphate; UDP: uridine diphosphate; TDP: thymidine diphosphate. over-expression of the UDP-glucose pyrophosphorylase gene and to be exported out of the cell membranes to validate their functions phosphoglucomutase gene was found to promote EPS production in such as protection against extreme environments. The secretion process both Streptococcus thermophilus and Bacillus licheniformis (Levander, requires a multi-component transport system for the export of carbo- Svensson, & Radstrom, 2002; Liu, Chen, Yang, Li, & He, 2017), in- hydrates with complex molecular structures. The intracellular poly- dicating single interventions in metabolic pathways may not be suffi- merization and transport of microbial EPSs mainly follow three me- cient to improve the production of desired EPSs, while multiple inter- chanisms (Fig. 3): (1) Wzx/Wzy-dependent pathway, (2) ATP-binding ventions are more likely to be efficient. However, with reference to cassette (ABC) transporter-dependent pathway, and (3) synthase-de- other studies, the increased enzyme specific activities for improvement pendent pathway (Ates, 2015). of UDP-sugar precursor availability is still controversial for guaran- The Wzx/Wzy-dependent pathway is considered as a major me- teeing higher EPS production (Boels, Ramos, Kleerebezem, & de Vos, chanism for Gram-negative bacteria to produce various EPSs. Most of 2001; Fialho et al., 2008). Extremophilic EPS production usually ac- the EPSs assembled by the Wzx/Wzy-dependent pathway are hetero- companies all the growth phases against the extreme environments, polysaccharides due to the presence of multiple GTs (Schmid et al., which can be different from the factors inducing mesophilic or neu- 2015). The assembled EPS is translocated across the cytoplasmic trophilic EPS production. It should be cautioned that over-expression of membrane by Wzx flippase and polymerized by Wzy polymerase. Fi- the genes specific for NDP-sugar synthesis could generate a metabolic nally, the EPS is transported by polysaccharide co-polymerase (PCP) burden for the growth of extremophilic strains. and outer membrane polysaccharide export (OPX) protein (Islam & For the second stage of EPS biosynthesis, glycosyltransferases Lam, 2014). This transport system was not considered as a major rate- transfer the activated nucleotide sugar precursors to the polysaccharide limiting step for EPS biosynthesis, instead the concomitant gene over- chain on a membrane-associated anchor for elongation. A significantly expression of the PCP and OPX proteins generated higher molecular increased EPS production without any deleterious effect to growth was weight EPS compared with that of the wild-type strain. Besides, it was achieved by over-expression of the gene encoding priming GT for also disclosed that the higher ratio of over-expressed PCP-OPX proteins linking the first galactose moiety to the lipid carrier in a mesophilic to Wzy polymerase could shift toward the polymerization of longer EPS strain Sinorhizobium meliloti (Jones, 2012). The combination of in- chains, and vice versa (Galván et al., 2013). creasing EPS precursor availability with engineered GTs in the EPS ABC transporter-dependent pathway uses ABC transporter to export biosynthetic route may synergistically enhance the production of ex- the EPS across inner membrane instead of Wzx and Wzy proteins, and tremophilic EPS. In addition, the metabolic control analysis can be the final secretion is still mediated by PCP and OPX proteins as Wzx/ performed for extremophilic EPS biosynthetic machinery to reveal the Wzy-dependent pathway (Whitney & Howell, 2013). ABC transporter- control points, in order to disclose the most efficient metabolic en- dependent pathway was considered as mainly presenting in capsular gineering strategy and combine it with highly activated GT system for polysaccharide biosynthesis (Schmid et al., 2015). However, thermo- elevated EPS production (Boels et al., 2001). philic strain Brevibacillus thermoruber 423 was suggested to be following After the assembly process, these hydrophilic macromolecules need ABC transporter-dependent pathway for its EPS transport due to the

18 J. Wang et al. Carbohydrate Polymers 205 (2019) 8–26

Fig. 3. Potential engineering targets for higher production and modified chain length during EPS assembly and transportation in ex- tremophiles. NDP: nucleoside diphosphate; GT: glycosyltransferase; Wzx: flippase; Wzy: poly- merase; PCP: polysaccharide co-polymerase; OPX: outer membrane polysaccharide export protein; ABC: ATP-binding cassette trans- porter; TPR: tetratricopeptide repeat protein; c- di-GMP: cyclic diguanylic acid.

presence of associated genes in its genome (Yildiz, Radchenkova, Arga, phenomenon indicates those acidophilic EPSs might be produced Kambourova, & Oner, 2015). The ABC transporter-dependent pathway through the synthase-dependent pathway. Intriguingly, besides acid- in this Gram-positive thermophilic bacterium diverged from that in ophilic EPSs, until now none of the other types of extremophilic EPSs well-studied mesophilic Gram-negative bacteria due to the appearance were discovered to be using the synthase-dependent pathway for as- of tetratricopeptide repeat (TPR) protein instead of PCP, indicating that sembly and secretion, or utilizing c-di-GMP as a stimulator to promote different tactics are required for further engineering of extremophilic EPS production. Therefore, it would be valuable to disclose if those EPS strains compared with mesophilic strains for improvement of EPS se- synthases could be more acidoresistant than the enzymes and trans- cretion. The ABC transporter might also be able to dictate the EPS chain porters within Wzx/Wzy- and ABC transporter-dependent pathway, and length (Schmid, 2018). Acquisition of high-resolution crystal structures why the EPSs synthesized through the synthase-dependent pathway of these proteins involved in extremophilic EPS polymerization and could provide protection against extreme acidic condition. transportation will be significant for generating mechanisms and pro- The biosynthesis of EPSs can also take place extracellularly through viding information for engineering strategies (Morgan, Strumillo, & the dissociated enzymes generated and secreted by the bacterial cells. Zimmer, 2012). This EPS synthetic process does not compete with cell growth for ac- In the synthase-dependent pathway, the repeating units of EPS are tivated monosaccharide precursors or lipid carriers (Prechtl et al., polymerized and then translocated by a synthase or synthase complex. 2018). Halophilic bacterium Halomonas smyrnensis AAD6T was able to Due to the reduced enzyme or enzyme system in this process compared synthesize levan as EPS through a secreted levansucrase. The over-ex- with other pathways, the molecular structure of EPSs synthesized pression of levansucrase could be attained by boric acid as a stimulator through the synthase-dependent pathway is simplified, and the syn- through the quorum sensing based signaling effect (Sarilmiser, Ates, thase-dependent pathway is quite favorable for biosynthesis of homo- Ozdemir, Arga, & Oner, 2015). Moreover, the gene encoding phos- polysaccharides or simple heteropolysaccharides with only two mono- phocarrier protein of the phosphoenolpyruvate sugar phospho- saccharide units (Schmid et al., 2015). Therefore, it can be speculated transferase system (PTS) for fructose uptake was knocked out in Halo- that the extremophilic EPSs with highly diverse monosaccharide com- monas smyrnensis AAD6T, and the mutant strain displayed an almost position could not be produced through the synthase-dependent threefold higher efficiency profile of levan production compared with pathway. For the mesophilic EPS synthesized through the synthase- the wild-type strain (Aydin, Ozer, Oner, & Arga, 2018). The supple- dependent mechanism, no correlation was observed between the mentation of mannitol to the culture medium also reduced the meta- number of synthase complexes and the EPS production level (Maleki, bolic requirement of fructose in Halomonas smyrnensis AAD6T since the Almaas, Zotchev, Valla, & Ertesvåg, 2016). Meanwhile, the higher ex- mannitol could be directly converted to fructose intracellularly (Ates pression of the genes encoding EPS synthase complex could link to et al., 2013). Both of these two strategies inhibited the uptake of increased molecular weight of the EPS (Díaz-Barrera, Soto, & fructose and thus accumulated more fructose moieties extracellularly as Altamirano, 2012). precursors for levan biosynthesis (Fig. 2). The cyclic diguanylic acid (c-di-GMP) is a bacterial secondary messenger enhancing the activity of EPS synthase with a c-di-GMP 3.2. Monosaccharide component modification strategies binding domain, and this regulation mechanism is significantly dif- ferent from other types of EPS synthetic pathways (Maleki et al., 2016; Up to the recent reports, the modification of monosaccharide units Morgan et al., 2012). The upregulation of c-di-GMP level can be a in the EPS backbone could be attained mainly using three different strategy to increase EPS production through synthase-dependent tactics, which pave the way for the production of tailor-made ex- pathway. The mutant with removal of the gene coding for tyrosine tremophilic EPSs. During the first phase within EPS production, the phosphatase which repressed the activity of diguanylate cyclase re- synthesis of a certain type of NDP-sugar precursor can be weakened to sponsible for c-di-GMP synthesis demonstrated 28-fold more EPS pro- change the monosaccharide contents in the EPS backbone. A mesophilic duction (Ueda & Wood, 2009). The downregulation of the activity of bacterium Paenibacillus elgii was identified containing two genes coding phosphodiesterase which degraded c-di-GMP could also be another way for uridine diphosphate-glucuronic acid (UDP-GlcA) decarboxylase, to increase the activity of EPS synthase (Hammer & Bassler, 2009). which could transfer UDP-GlcA to UDP-xylose. The single-gene The c-di-GMP has been found as an activator for EPS biosynthesis by knockout mutant of UDP-GlcA decarboxylase produced the EPS with several acidophilic bacteria including Acidithiobacillus species and higher glucuronic acid and lower xylose content compared with that of Leptospirillum ferriphilum (Christel et al., 2018; Díaz, Castro, Copaja, & the wild-type strain. Meanwhile no significant variation in mannose Guiliani, 2018; Ruiz, Castro, Barriga, Jerez, & Guiliani, 2011). This and glucose content was observed between the EPSs produced by the

19 J. Wang et al. Carbohydrate Polymers 205 (2019) 8–26 single-gene knockout mutant and wild-type strain (Li et al., 2015). and further study for EPS characterization and transcriptomics is highly Another genetic engineering strategy targets the genes coding for recommended to elucidate the EPS biosynthetic and regulatory me- glycosyltransferases responsible for monomer assembly. The condi- chanism in this psychrophilic bacterium. tional reduction of the activity of the priming GT involved in attaching the first monosaccharide unit to the lipid carrier in mesophilic bac- 4. Recent progress in the application of extremophilic EPSs terium Lactobacillus rhamnosus could decrease the amount of repeating unit modules available for polymerase, leading to premature chain 4.1. Biomedical application termination; thus, shorter EPS chains were secreted (Bouazzaoui & LaPointe, 2006). The removal of gene encoding a non-priming GT was 4.1.1. Antitumor and immunoregulatory effect able to block the addition of the undesired monosaccharide unit onto Currently, accumulated evidence has demonstrated that ex- the EPS polymer chain. In a current study concerning the EPS biosyn- tremophilic EPSs have a broad spectrum of biological activities, such as thetic machinery in a mesophilic strain Paenibacillus polymyxa, the anti-cancer, anti-oxidant and immunoregulatory properties, which can state-of-the-art CRISPR-Cas9 genome editing tool was applied for gene be promising for biomedical applications. The anti-cancer efficacy has knockout strategy in order to modify its EPS monomer composition. already been recognized for the polysaccharides generated by fungi, The EPS variants with altered monosaccharide distribution and rheo- algae, and plants (Zong, Cao, & Wang, 2012). Extremophilic EPSs can logical behavior from the wild-type EPS were obtained by disruption of hardly be highly cytotoxic against malignant proliferating mammalian the gene coding for one of the non-priming glycosyltransferases within cells like chemotherapeutic drugs. Instead, extremophilic EPSs may its EPS biosynthetic system (Rütering et al., 2017). induce apoptosis in tumor cells via coupling specific surface receptors The heterologous expression of the exopolysaccharide gene cluster (Ruiz-Ruiz et al., 2011). The thermophilic EPSs from Geobacillus sp. to the recombinant strain also leads to generation of the EPS with dif- TS3-9 was found to significantly inhibit the proliferation of hepatoma ferent monosaccharide composition from the native strain. The gene carcinoma cell in a dose-dependent manner in vitro (Wang et al., 2017). cluster with functional regions coding for EPS biosynthesis regulatory Further study of antiproliferation effects on non-tumor cells is required protein, glycosyltransferases, EPS chain-length determinator, poly- to identify its antitumor specificity. Furthermore, the addition of non- merase and transporter from a thermotolerant bacterium Streptococcus sugar functional groups onto extremophilic EPS molecules may gen- thermophilus Sfi6 was expressed heterologously in a non-EPS-producing erate improved biological activities (e.g. antitumor activity) compared strain Lactococcus lactis MG1363. The transferred EPS biosynthetic with native EPSs. An oversulphated EPS produced by a halophilic system could utilize the NDP-sugars generated in the host strain bacterium Halomonas stenophila B100 could specifically induce apop- through its house-keeping genes as building blocks for EPS production. tosis of leukemia cells from peripheral blood, and the addition of sul- For the recombinant EPS, the N-acetylgalactosamine (GalNAc) moieties phate moieties to native EPS was considered to enhance its anti- in the backbone was replaced by galactose residues, since the host proliferative efficacy (Ruiz-Ruiz et al., 2011). Another halophilic EPS strain was unable to synthesize the corresponding UDP-GalNAc pre- levan was modified through periodate oxidation to harbor aldehyde cursor. Meanwhile, the recombinant EPS also lacked the galactose side- groups, and the aldehyde-activated levan derivatives showed both chain compared with the wild-type EPS (Stingele et al., 1999). This biocompatibility to non-tumor cells and anti-cancer activity against heterologous strategy requires that the polymerases and transporters several human tumor cell lines in vitro. The antitumor efficacy was from the native strain can recognize the recombinant EPS chain without confirmed to be enhanced by increasing the oxidation degree of the EPS strong exclusive selectivity. Many extremophilic bacteria are able to (Sarilmiser & Oner, 2014). generate two or three EPSs with various molecular weight and EPSs can also inhibit tumor progression through the im- monomer distributions (Table 1). Therefore, the polymerization and munoenhancement effect (Zong et al., 2012). The EPS from a thermo- export system from the extremophiles with multi-EPS producing cap- philic bacterium Thermus aquaticus YT-1 was proved to be an im- ability may not be strictly specific towards a single type of EPS back- munomodulator which stimulated macrophage cells to produce the bone, and this infidelity provides a further degree of flexibility in cytokines tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6). producing recombinant extremophilic EPS in an engineered strain This thermophilic EPS also induced macrophages to release nitric oxide through heterologous genetic engineering techniques. (NO) as inflammatory mediator. Toll-like receptor 2 (TLR2) expressed on the immune cell surface was confirmed as the natural receptor of this 3.3. Genome annotation for extremophilic EPS biosynthetic system thermophilic EPS (Lin et al., 2011). A psychrophilic EPS produced by Pseudoalteromonas sp. strain S-5 showed a similar stimulation effect on The genome annotation for extremophilic strains can be a powerful macrophages to secrete TNF-α and NO. Meanwhile, it could also en- tool to disclose the essential genes associated with EPS biosynthesis and hance the phagocyte function of macrophage cells (Bai et al., 2012). hence gain more insight about the biological mechanisms of EPS pro- The stimulation of the production of TNF-α and NO indicates a role of duction, serving as the starting point to develop genetic and metabolic these EPSs in activation of macrophages into the M1 subtype (classi- engineering strategies to optimize EPS production and modify EPS cally activated macrophages), which can suppress tumor growth, me- monomer composition to fit industrial and biomedical requirements. tastasis and angiogenesis (Chen et al., 2018). Thus, these extremophilic Currently, a preliminary model of the EPS biosynthesis mechanism was EPSs have high potential for application in macrophage-mediated im- successfully proposed for a thermophilic bacterium Brevibacillus ther- mune therapy for cancer treatment. moruber 423 through whole-genome analysis (Yildiz et al., 2015). The Several extremophilic EPSs were found to be non-cytotoxic for genome annotation of halophilic strain Halomonas smyrnensis AAD6T normal cell lines. The heteropolysaccharide produced by thermophilic revealed the presence of a Pel exopolysaccharide gene cluster in its bacterium Brevibacillus thermoruber strain 423 demonstrated high bio- genome, indicating its capacity to produce Pel EPS besides being a compatibility to a monkey kidney fibroblast cell line (Yildiz et al., levan producer (Diken et al., 2015). The genome analysis was also 2014). Levan secreted by halophile Halomonas smyrnensis AAD6T also carried for acidophilic bacterium Leptospirillum ferriphilum, which de- showed high biocompatibility and affinity with non-cancerous cell monstrated cellulose and Pel EPS synthetic genes involved in the syn- lines. The lack of branch structure in this halophilic levan molecule was thase-dependent pathway (Christel et al., 2018). With a relatively considered as crucial for the absence of cytotoxic activity (Küçükaşik larger genome size, the psychrophile Phormidesmis priestleyi BC1401 et al., 2011; Poli et al., 2009). The EPSs from halothermophilic bacteria contained both gene clusters following the scheme of Wzx/Wzy-de- Bacillus licheniformis strain T14 and B3-15 were non-cytotoxic toward pendent and ABC transporter-dependent EPS export systems (Chrismas, human peripheral blood mononuclear cells (PMBC) at maximum con- Barker, Anesio, & Sánchez-Baracaldo, 2016). This is quite intriguing, centration of 400 and 300 μg/ml, respectively (Arena et al., 2006;

20 J. Wang et al. Carbohydrate Polymers 205 (2019) 8–26

Gugliandolo, Spanò, Lentini, Arena, & Maugeri, 2013). Additionally, important property of bioemulsifiers from extremophiles is their high thermophilic EPS synthesized by Geobacillus thermodenitrificans strain emulsion stability over a wide range of temperature, pH and salinity B3-72 was also non-toxic to PMBC cells at concentration of 300 μg/ml (Zheng et al., 2012). For example, in Arias’ report, neither the viscosity or below (Arena et al., 2009). Psychrophilic EPS of Pseudoalteromonas nor the pseudoplasticity of mauran (a halophilic EPS) solution was af- sp. S-5 had no cytotoxic effect under 500 μg/ml against murine peri- fected by the presence of salts, sugar, surfactants, lactic acid, changes in toneal macrophages (Bai et al., 2012). These non-cytotoxic ex- pH, or freezing and thawing (Arias et al., 2003). The minor content of tremophilic EPSs are highly promising for application as a biocompa- protein in those extremophilic EPSs might be essential for the emulsi- tible carrier to conjugate antineoplastic drug and targeting ligand such fying activity (Llamas, Amjres, Mata, Quesada, & Béjar, 2012). The as monoclonal antibody. This three-phase antitumor drug conjugate presence of uronic acid and acetyl group in EPS was also considered to would attain the specific delivery of non-selective cytotoxic drugs to contribute to its emulsifying capacity (Caruso et al., 2017; Jain, Mody, tumor tissue with enhanced antitumor efficacy and selectivity Mishra, & Jha, 2012; Mata et al., 2006). (Asamoah-Asare, Zhang, & Chen, 2013; Dragojevic, Ryu, & Raucher, Pseudoplastic rheological behavior is another common property 2015). The biodegradability of these extremophilic EPSs must be among those extremophilic EPSs with emulsifying activity (Table 1). identified in order to make in vivo test and future clinical trial feasible. High pseudoplasticity is an attractive rheological characteristic in di- The immunoregulatory effect of several extremophilic EPSs also verse types of food formulations, such as sauce, dairy, cake, salad leads to antiviral activity. The EPSs from halothermophilic strain dressing, syrup, and pudding (Bahram Parvar & Razavi Seyed, 2012; Bacillus licheniformis strain B3-15, Bacillus licheniformis strain T14, and Han et al., 2014). For the development of foodstuffs, the pseudoplastic thermophile Geobacillus thermodenitrificans strain B3-72 decreased property of EPS is advantageous to generate comfortable sensory herpes simplex virus type 2 (HSV-2) replication in PMBC through sti- properties such as mouth feel and flavor release. It is also useful for food mulating the expression of different proinflammatory cytokines in- processes, such as mixing, pouring and pumping with different opera- volved in the immune surveillance toward virus infection, indicating a tive shear rates (Han et al., 2014). potential application as therapy in herpes virus infection and im- Based on current extremophilic EPS research, it remains difficult for munocompromised host (Arena et al., 2006, 2009; Spanò & Arena, the newly discovered extremophilic EPSs to displace the commercia- 2016). lized biopolymers such as xanthan and gellan in the food industry, due to the costly production processes of extremophilic EPSs using pure 4.1.2. Antioxidant effect sugar as substrate. Inexpensive and renewable substrates need to be Antioxidant activity leads to scavenging reactive oxygen species used for EPS production, such as molasses which can be a feasible (ROS), which generate oxidative stress to neuronal cells and are deeply substitute for sucrose (Küçükaşik et al., 2011; Sam et al., 2011). A associated with chronic and degenerative diseases such as neurode- continuous cultivation technique with economic potential for industrial generative disorders (Xu, Bi, & Wan, 2016). Polysaccharides have been scale production also showed a much higher efficiency for the fer- shown to play a crucial role as natural antioxidants for the prevention mentation of extremophilic EPS in comparison with batch cultures of oxidative damage in the human body (Wang et al., 2013). Ex- (Radchenkova et al., 2015). tremophilic EPSs are usually non-pathogenic and their high biodiversity offers various biotechnological activities including antioxidancy. Ha- 4.3. Biomaterial application lophilic EPS isolated from Halolactibacillus miurensis, and thermophilic EPS from Geobacillus sp. strain TS3-9 both demonstrated dose-depen- 4.3.1. EPS-based nanoparticles dent scavenging activity against DPPH (2,2-diphenyl-1-picrylhydrazyl), Nanotechnology is an indispensable discipline in the modern hydroxyl and superoxide free radicals (Arun et al., 2017; Wang et al., pharmaceutical field for a variety of applications. The non-ideal bio- 2017). Additionally, the EPS produced by halothermophilic bacterium degradability and biocompatibility of the synthetic nanoparticles Halomonas nitroreducens WB1 had antioxidant properties to scavenge usually impair renal excretion and induce many unacceptable side ef- hydroxyl and DPPH radicals (Chikkanna, Ghosh, & Kishore, 2018). A fects. During the constant lookout for novel products with improved hyper-branched psychrophilic EPS secreted by Polaribacter sp. SM1127 pharmaceutical functions, the extremophilic EPSs have received in- showed substantially higher antioxidant activity than that of hyaluronic creasing attention as alternatives to synthetic polymers in the produc- acid, an industrial annexing agent for scavenging radicals (Sun et al., tion and modification of nanoparticles. Their valuable biological char- 2015). These extremophilic EPSs with remarkable antioxidative capa- acteristics and simplicity of chemical modifications make them strong city may be efficacious in the treatment of neurodegenerative diseases. candidates for nanoparticle application (Raveendran et al., 2014). Furthermore, they can be studied as natural dietary antioxidants for the Extremophilic EPSs can be applied to nanoparticle technology in inactivation of oxyradicals and reduction of the incidence rate of neu- two ways: one is to utilize EPS to form nanoparticles directly, and the rodegenerative disease. other is to use EPS to encapsulate nanoparticles made from another material. The extremophilic EPSs with negatively charged groups could 4.2. Food application perform as polyelectrolytes, which allow them to self-assemble with positively charged biomolecules to form biodegradable nanoparticles The microbial polysaccharides which are able to stabilize emulsions through polyelectrolyte complexation. As a positively charged biopo- between water and hydrophobic compounds have potential as natural lymer, is often used for polyelectrolyte nanoparticle formation emulsifiers in the food industry (Freitas et al., 2009). Bioemulsifiers with negatively charged EPSs (Deepak, Pandian, Sivasubramaniam, have the advantages of biodegradability, low toxicity, selectivity and Nellaiah, & Sundar, 2015; Karlapudi et al., 2016). Halophilic EPS environmental compatibility over artificial products (Mata et al., 2008). mauran-chitosan hybrid nanoparticles, produced through an ionic-ge- The emulsifying activity has been found to be common in extremophilic lation method, manifested stable drug release and biocompatibility EPSs, including those produced by thermophiles, psychrophiles, halo- when used for antitumor drug encapsulation (Raveendran, Poulose philes, and alkaliphiles; and they are all heteropolysaccharides et al., 2013). Another halophilic EPS levan, produced by Halomonas (Table 1). Among those extremophilic EPSs, EPS from psychrophilic smyrnensis AAD6T, was investigated for nanoparticle formation through bacterium Pseudomonas sp. ID1 had a higher emulsifying activity than self-assembly, and its suitability was affirmed as a nanocarrier for de- xanthan gum and arabic gum for several food oils (Carrión, Delgado, & livery of peptides and proteins (Sezer, Kazak, Öner, & Akbuğa, 2011). Mercade, 2015). Application of EPSs in the coating or stabilization of chemically In industrial processes, emulsifiers may be exposed to extremes of synthetic nanoparticles also has important potential. Some synthetic temperature, environmental pH and salinity (Freitas et al., 2009). An nanoparticles possess irreplaceable properties, such as a

21 J. Wang et al. Carbohydrate Polymers 205 (2019) 8–26 photoluminescence for imaging, but their toxicity in the human body solubility of EPSs avoids using toxic solvents or additives during elec- needs to be minimized through suitable modification. Polysaccharide trospinning process, making EPS-based electrospun materials excellent coated nanoparticles have already been shown to possess rapid uptake candidates for biomedical engineering applications. However, it can be and internalization through the endocytosis effect compared with un- difficult to generate neat EPS fibers by electrospinning, since EPS so- coated nanoparticles, and the cellular toxicity of EPS-coated nano- lutions tend to have high surface tension, non-ideal viscosity, and ex- particles was also notably reduced (Banerjee & Bandopadhyay, 2016). cessively strong charge density due to the anionic nature of EPSs Extremophilic EPSs can therefore be applied as a passivation agent to (Santos et al., 2014; Torres-Giner et al., 2008). The addition of a hy- improve the biocompatibility of nanoparticles, and make them more drophilic co-polymer as a carrier agent is one way to circumvent the feasible for pharmaceutical applications (Raveendran et al., 2014). For limitations from those bio-polyelectrolyte (e.g. EPS) solutions instance, quantum dots (QDs) are nanocrystals with a photoluminescent (Vashisth, Pruthi, Singh, & Pruthi, 2014). For example, polyvinyl al- property and applied as preferred imaging agents in biological tissues cohol (PVA), a biocompatible and water-soluble polymer, can be for clinical diagnose (Deepagan et al., 2012; Raveendran et al., 2014). blended with EPS, reducing the repulsive force from the negatively However, QDs such as ZnS nanocrystals are synthesized from toxic charged EPS solution and allowing the generation of uniform nanofi- chemicals in order to maintain their imaging property. These types of bers by electrospinning (Qian et al., 2016; Santos et al., 2014; Vashisth QDs are hydrophobic and water-insoluble, which hinder their applica- et al., 2014). The halophilic EPS mauran was blended with PVA and tion in the medical field. One solution to overcome these drawbacks is electrospun to generate a scaffold with continuous, uniform nanofibers. to stabilize QDs with a capping agent, and extremophilic EPSs can be a The mauran-based nanofiber was able to boost cellular adhesion, mi- stabilizing agent for QDs to improve their cellular acceptance. In a gration, proliferation, and differentiation of mammalian cells in vitro. study, the stabilization of ZnS-Mn QDs using halophilic EPS mauran The polyanionic nature of extremophilic EPSs increases the negative was highly successful in imparting a biocompatible and safe mode of charge accumulation on the surface of the scaffold, which is helpful for cellular imaging under in vitro conditions. Anionic EPSs are able to bind protein adsorption and the ability to enhance cellular attachment. An with nanoparticles having positive charge. Additionally, the acetyl excellent property of mauran is that it can keep the same viscosity groups present in EPSs can bring more positively charged ions to the under a high concentration of salt and sugar, or under extreme pH vicinity of binding sites, thus allowing stronger binding (Raveendran values; and the stable viscosity of a mauran solution is highly ad- et al., 2014). vantageous in obtaining stable electrospinning conditions (Raveendran, Dhandayuthapani et al., 2013). In future studies, those extremophilic 4.3.2. EPS-based films EPSs with heavy metal adsorption capability may also be blended with Adhesive and biocompatible films are an attractive way to offer PVA and then electrospun onto a basal microfiltration membrane for fixation to tissues either externally, for wound healing, or internally as water filtration applications (Santos et al., 2014). a surgical sealant. For the purpose of medical adhesives, such as a burn dressing, drug delivery, and implantation, the films are required to keep good long-term performance on skin or in biological fluids without 5. Conclusion and prospective triggering a pathological process (Costa et al., 2013). The inherent functions of extremophilic EPSs are to provide adhesion and protection It is now widely considered that extremophilic microorganisms to bacteria in an extreme environment. Therefore, naturally derived provide a valuable resource, not only for the elucidation of bioprocesses EPS-based films confer sufficient cohesive strength and maintain a in extreme environments, but also for the exopolysaccharides they biocompatible response to cells and tissues. The negative charge allows produce, which have a valuable range of physicochemical properties EPSs to be adsorbed by electrostatic self-assembly and sequential for- and highly promising commercial applications. To realize the full value mation onto multilayer film. The EPS levan produced by halophilic of these biopolymers, it will be necessary to gain insight into the bacterium Halomonas smyrnensis AAD6T was applied by electrostatic modulation of EPS biosynthesis in extremophiles. The genome anno- adsorption to construct multilayer film, which demonstrated a pro- tation and construction of EPS biosynthetic pathways will be of sig- mising enhancement of live cell adhesive property. The extremophilic nificant importance in determining the kind of monosaccharide units EPS-based film surface, having better biocompatibility, provides a va- that can be incorporated into the structures of the EPSs, and how they luable means to explore novel types of cell-material interactions, are incorporated; as well as how the compositions of monosaccharides leading to an understanding of how to promote or inhibit specific cel- in the EPSs can be affected. Metabolic and genetic engineering will lular responses when contacting bio-based materials (Costa et al., enable the development of effective strategies to successfully enhance 2013). Moreover, the halophilic EPS levan was mixed with chitosan and EPS production and engineer EPS properties via tailoring their chemical polyethylene oxide (PEO) to produce hybrid films through a solvent composition and structure. Applications of extremophilic EPSs in bio- casting method. This ternary blend film showed better biocompatible medical, food and pollution-mitigation products can be foreseen based behavior compared with chitosan-PEO binary film (Bostan et al., 2014). on their known biodegradability, biocompatibility, non-toxicity, and Another method applied for thin film production using halophilic EPS chemical functionality. It can be anticipated that more extremophilic levan was matrix-assisted pulsed laser evaporation (MAPLE). This na- EPSs from different harsh environments will be discovered, character- nostructured film was also able to sustain cell adhesion and prolifera- ized, and optimized via evolving bioengineering protocols, and that tion (Sima et al., 2011). they will be employed in high value-added industries with potential for strong, sustainable growth. 4.3.3. EPS-based materials through electrospinning Electrospinning is a versatile and relatively cost-effective technique to fabricate a large variety of soluble or fusible synthetic and natural Acknowledgements polymers into continuous fibers with diameters in the submicron to nanometer range (Salem, 2007). The electrospinning method has al- This research was supported by the National Science Foundation in ready been applied in many technological fields, and it enables the the form of BuG ReMeDEE initiative (Award # 1736255). Authors also production of novel biomaterials using naturally occurring biopolymers acknowledge the financial support in the form of CNAM/Bio Centre with complex molecular structures (Torres-Giner, Ocio, & Lagaron, provided by the South Dakota Governor’sOffice of Economic 2008). Due to the natural properties of extremophilic EPSs, the mate- Development. Research support from the Department of Chemical and rials electrospun from those EPSs are considered sustainable, bio- Biological Engineering at the South Dakota School of Mines and compatible, biodegradable, and non-toxic. Additionally, the high water- Technology is gratefully acknowledged.

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References Chen, M., Li, Y., Liu, Z., Qu, Y., Zhang, H., Li, D., ... Liu, M. (2018). Exopolysaccharides from a Codonopsis pilosula endophyte activate macrophages and inhibit cancer cell proliferation and migration. Thoracic Cancer, 9(5), 630–639. Arena, A., Gugliandolo, C., Stassi, G., Pavone, B., Iannello, D., Bisignano, G., ... Maugeri, Chikkanna, A., Ghosh, D., & Kishore, A. (2018). Expression and characterization of a fi T. L. (2009). An exopolysaccharide produced by Geobacillus thermodenitri cans strain potential exopolysaccharide from a newly isolated halophilic thermotolerant bacteria B3-72: Antiviral activity on immunocompetent cells. Immunology Letters, 123(2), Halomonas nitroreducens strain WB1. PeerJ, 6, e4684. – 132 137. Chrismas, N. A. M., Barker, G., Anesio, A. M., & Sánchez-Baracaldo, P. (2016). Genomic Arena, A., Maugeri, T. L., Pavone, B., Iannello, D., Gugliandolo, C., & Bisignano, G. mechanisms for cold tolerance and production of exopolysaccharides in the Arctic ff (2006). Antiviral and immunoregulatory e ect of a novel exopolysaccharide from a cyanobacterium Phormidesmis priestleyi BC1401. BMC Genomics, 17(1), 533. marine thermotolerant Bacillus licheniformis. International Immunopharmacology, 6(1), Christel, S., Herold, M., Bellenberg, S., El Hajjami, M., Buetti-Dinh, A., Pivkin, I. V., ... – 8 13. Dopson, M. (2018). Multi-omics reveals the lifestyle of the acidophilic, mineral-oxi- Arias, S., del Moral, A., Ferrer, M. R., Tallon, R., Quesada, E., & Bejar, V. (2003). Mauran, dizing model species Leptospirillum ferriphilumT. Applied and environmental micro- an exopolysaccharide produced by the halophilic bacterium Halomonas maura, with a biology, 84(3), e02091–02017. novel composition and interesting properties for biotechnology. Extremophiles, 7(4), Christensen, B. E., Kjosbakken, J., & Smidsrød, O. (1985). Partial chemical and physical – 319 326. characterization of two extracellular polysaccharides produced by marine, periphytic Arun, J., Selvakumar, S., Sathishkumar, R., Moovendhan, M., Ananthan, G., Maruthiah, Pseudomonas sp. Strain NCMB 2021. Applied and environmental microbiology, 50(4), T., ... Palavesam, A. (2017). In vitro antioxidant activities of an exopolysaccharide 837–845. from a salt pan bacterium Halolactibacillus miurensis. Carbohydrate Polymers, 155, Corsaro, M. M., Grant, W. D., Grant, S., Marciano, C. E., & Parrilli, M. (1999). Structure – 400 406. determination of an exopolysaccharide from an alkaliphilic bacterium closely related Asamoah-Asare, J., Zhang, Y., & Chen, Y. (2013). Antibody conjugated polymeric pro- to Bacillus spp. European Journal of Biochemistry, 264(2), 554–561. drugs the future for cancer therapy. International Journal of Advanced Biotechnology Corsaro, M. M., Lanzetta, R., Parrilli, E., Parrilli, M., Tutino, M. L., & Ummarino, S. – and Bioengineering, 1(1), 1 17. (2004). Influence of growth temperature on lipid and phosphate contents of surface Ates, O. (2015). Systems biology of microbial exopolysaccharides production. Frontiers in polysaccharides from the Antarctic bacterium Pseudoalteromonas haloplanktis TAC Bioengineering and Biotechnology, 3, 200. 125. Journal of bacteriology, 186(1), 29–34. ff Ates, O., Arga, K. Y., & Oner, E. T. (2013). The stimulatory e ect of mannitol on levan Costa, R. R., Neto, A. I., Calgeris, I., Correia, C. R., Pinho, A. C. M., Fonseca, J., ... Mano, J. biosynthesis: Lessons from metabolic systems analysis of Halomonas smyrnensis F. (2013). Adhesive nanostructured multilayer films using a bacterial exopoly- – AAD6T. Biotechnology Progress, 29(6), 1386 1397. saccharide for biomedical applications. Journal of Materials Chemistry B, 1(18), Aydin, B., Ozer, T., Oner, E. T., & Arga, K. Y. (2018). The genome-based metabolic sys- 2367–2374. tems engineering to boost levan production in a halophilic bacterial model. OMICS: A d’Hugues, P., Joulian, C., Spolaore, P., Michel, C., Garrido, F., & Morin, D. (2008). – Journal of Integrative Biology, 22(3), 198 209. Continuous bioleaching of a pyrite concentrate in stirred reactors: Population dy- Bahram Parvar, M., & Razavi Seyed, M. A. (2012). Rheological interactions of selected namics and exopolysaccharide production vs. bioleaching performance. – – – fi hydrocolloid sugar milk emulsi er systems. International Journal of Food Science & Hydrometallurgy, 94(1-4), 34–41. – Technology, 47(4), 854 860. Deepagan, V. G., Sarmento, B., Menon, D., Nascimento, A., Jayasree, A., Bai, Y., Zhang, P., Chen, G., Cao, J., Huang, T., & Chen, K. (2012). Macrophage im- Sreeranganathan, M., ... Rangasamy, J. (2012). In vitro targeted imaging and delivery munomodulatory activity of extracellular polysaccharide (PEP) of Antarctic bac- of camptothecin using cetuximab-conjugated multifunctional PLGA-ZnS nano- – terium Pseudoaltermonas sp.S-5. International Immunopharmacology, 12(4), 611 617. particles. Nanomedicine, 7(4), 507–519. Baker-Austin, C., & Dopson, M. (2007). Life in acid: pH homeostasis in acidophiles. Trends Deepak, V., Pandian, S. R. K., Sivasubramaniam, S. D., Nellaiah, H., & Sundar, K. (2015). – in Microbiology, 15(4), 165 171. Synthesis of polyelectrolyte nanoparticles from anticancer exopolysaccharide isolated Banerjee, A., & Bandopadhyay, R. (2016). Use of dextran nanoparticle: A paradigm shift from probiotic Lactobacillus acidophilus. Research Journal of Microbiology, 10(5), in bacterial exopolysaccharide based biomedical applications. International Journal of 193–204. – Biological Macromolecules, 87, 295 301. Díaz, M., Castro, M., Copaja, S., & Guiliani, N. (2018). Biofilm formation by the acid- ff Bejar, V., Calvo, C., Moliz, J., Diaz-Martinez, F., & Quesada, E. (1996). E ect of growth ophile bacterium Acidithiobacillus thiooxidans involves c-di-GMP pathway and Pel conditions on the rheological properties and chemical composition of Volcaniella exopolysaccharide. Genes, 9(2), 113. – eurihalina exopolysaccharide. Applied Biochemistry and Biotechnology, 59(1), 77 86. Díaz-Barrera, A., Soto, E., & Altamirano, C. (2012). Alginate production and alg8 gene Béjar, V., Llamas, I., Calvo, C., & Quesada, E. (1998). Characterization of exopoly- expression by Azotobacter vinelandii in continuous cultures. Journal of Industrial saccharides produced by 19 halophilic strains of the species Halomonas eurihalina. Microbiology & Biotechnology, 39(4), 613–621. – Journal of Biotechnology, 61(2), 135 141. Diken, E., Ozer, T., Arikan, M., Emrence, Z., Oner, E. T., Ustek, D., ... Arga, K. Y. (2015). ff Bhalla, A., Bansal, N., Kumar, S., Bischo , K. M., & Sani, R. K. (2013). Improved lig- Genomic analysis reveals the biotechnological and industrial potential of levan pro- nocellulose conversion to biofuels with thermophilic bacteria and thermostable en- ducing halophilic extremophile, Halomonas smyrnensis AAD6T. SpringerPlus, 4(1), – zymes. Bioresource Technology, 128, 751 759. 393. Boels, I. C., Ramos, A., Kleerebezem, M., & de Vos, W. M. (2001). Functional analysis of Donot, F., Fontana, A., Baccou, J. C., & Schorr-Galindo, S. (2012). Microbial exopoly- the Lactococcus lactis galU and galE genes and their impact on sugar nucleotide and saccharides: Main examples of synthesis, excretion, genetics and extraction. exopolysaccharide biosynthesis. Applied and environmental microbiology, 67(7), Carbohydrate Polymers, 87(2), 951–962. – 3033 3040. Dopson, M., Baker-Austin, C., Koppineedi, P. R., & Bond, P. L. (2003). Growth in sulfidic Bostan, M. S., Mutlu, E. C., Kazak, H., Sinan Keskin, S., Oner, E. T., & Eroglu, M. S. (2014). mineral environments: Metal resistance mechanisms in acidophilic micro-organisms. fi Comprehensive characterization of chitosan/PEO/levan ternary blend lms. Microbiology, 149(Pt 8), 1959–1970. – Carbohydrate Polymers, 102, 993 1000. Dragojevic, S., Ryu, S. J., & Raucher, D. (2015). Polymer-based prodrugs: Improving Bouazzaoui, K., & LaPointe, G. (2006). Use of antisense RNA to modulate glycosyl- tumor targeting and the solubility of small molecule drugs in cancer therapy. transferase gene expression and exopolysaccharide molecular mass in Lactobacillus Molecules, 20(12), 21750–21769. – rhamnosus. Journal of Microbiological Methods, 65(2), 216 225. Drouillard, S., Jeacomine, I., Buon, L., Boisset, C., Courtois, A., Thollas, B., ... Helbert, W. Brian-Jaisson, F., Molmeret, M., Fahs, A., Guentas-Dombrowsky, L., Culioli, G., Blache, Y., (2015). Structure of an amino acid-decorated exopolysaccharide secreted by a Vibrio fi ... Ortalo-Magné, A. (2016). Characterization and anti-bio lm activity of extracellular alginolyticus strain. Marine Drugs, 13(11), 6723–6739. fi polymeric substances produced by the marine bio lm-forming bacterium Duckworth, A., Grant, W., Jones, B., & Van Steenbergen, R. (1996). Phylogenetic diversity – Pseudoalteromonas ulvae strain TC14. Biofouling, 32(5), 547 560. of soda lake alkaliphiles. FEMS microbiology ecology, 19(3), 181–191. Carrión, O., Delgado, L., & Mercade, E. (2015). New emulsifying and cryoprotective Ewert, M., & Deming, J. W. (2013). Sea ice microorganisms: Environmental constraints exopolysaccharide from Antarctic Pseudomonas sp. ID1. Carbohydrate Polymers, 117, and extracellular responses. Biology, 2(2), 603–628. – 1028 1034. Fialho, A. M., Moreira, L. M., Granja, A. T., Popescu, A. O., Hoffmann, K., & Sa-Correia, I. Caruso, C., Rizzo, C., Mangano, S., Poli, A., Di Donato, P., Finore, I., ... Giudice, A. L. (2008). Occurrence, production, and applications of gellan: Current state and per- (2017). Production and biotechnological potentialities of extracellular polymeric spectives. Applied Microbiology and Biotechnology, 79(6), 889–900. substances from sponge-associated Antarctic bacteria. Applied and environmental mi- Flemming, H. C., & Wingender, J. (2010). The biofilm matrix. Nature Reviews – crobiology, 84(4), e01624 01617. Microbiology, 8(9), 623–633. Caruso, C., Rizzo, C., Mangano, S., Poli, A., Di Donato, P., Nicolaus, B., ... Lo Giudice, A. Freitas, F., Alves, V. D., Carvalheira, M., Costa, N., Oliveira, R., & Reis, M. A. M. (2009). (2018). Extracellular polymeric substances with metal adsorption capacity produced Emulsifying behaviour and rheological properties of the extracellular polysaccharide by Pseudoalteromonas sp. MER144 from Antarctic seawater. Environmental Science and produced by Pseudomonas oleovorans grown on glycerol byproduct. Carbohydrate – Pollution Research, 25(5), 4667 4677. Polymers, 78(3), 549–556. Casillo, A., Parrilli, E., Sannino, F., Mitchell, D. E., Gibson, M. I., Marino, G., ... Corsaro, Freitas, F., Alves, V. D., & Reis, M. A. (2011). Advances in bacterial exopolysaccharides: M. M. (2017). Structure-activity relationship of the exopolysaccharide from a psy- From production to biotechnological applications. Trends in Biotechnology, 29(8), chrophilic bacterium: A strategy for cryoprotection. Carbohydrate Polymers, 156, 388–398. – 364 371. Galván, E. M., Ielmini, M. V., Patel, Y. N., Bianco, M. I., Franceschini, E. A., Schneider, J. Casillo, A., Ståhle, J., Parrilli, E., Sannino, F., Mitchell, D. E., Pieretti, G., ... Corsaro, M. M. C., ... Ielpi, L. (2013). Xanthan chain length is modulated by increasing the avail- (2017). Structural characterization of an all-aminosugar-containing capsular poly- ability of the polysaccharide copolymerase protein GumC and the outer membrane saccharide from Colwellia psychrerythraea 34H. Antonie van Leeuwenhoek, 110(11), polysaccharide export protein GumB. Glycobiology, 23(2), 259–272. – 1377 1387. Gehrke, T., Telegdi, J., Thierry, D., & Sand, W. (1998). Importance of extracellular Chawla, P. R., Bajaj, I. B., Survase, S. A., & Singhal, R. S. (2009). Microbial cellulose: polymeric substances from Thiobacillus ferrooxidans for bioleaching. Applied and en- Fermentative production and applications. Food Technology & Biotechnology, 47(2), vironmental microbiology, 64(7), 2743–2747. – 107 124. Gu, D., Jiao, Y., Wu, J., Liu, Z., & Chen, Q. (2017). Optimization of EPS production and

23 J. Wang et al. Carbohydrate Polymers 205 (2019) 8–26

characterization by a halophilic bacterium, Kocuria rosea ZJUQH from Chaka Salt 13(1), 71–77. Lake with response surface methodology. Molecules, 22(5), 814. Li, W., Zhou, W., Zhang, Y., Wang, J., & Zhu, X. (2008). Flocculation behavior and me- Guezennec, J. (2002). Deep-sea hydrothermal vents: A new source of innovative bacterial chanism of an exopolysaccharide from the deep-sea psychrophilic bacterium exopolysaccharides of biotechnological interest? Journal of Industrial Microbiology and Pseudoalteromonas sp. SM9913. Bioresource Technology, 99(15), 6893–6899. Biotechnology, 29(4), 204–208. Lin, L., & Xu, J. (2013). Dissecting and engineering metabolic and regulatory networks of Gugliandolo, C., Lentini, V., Spanò, A., & Maugeri, T. L. (2012). New bacilli from shallow thermophilic bacteria for biofuel production. Biotechnology Advances, 31(6), hydrothermal vents of Panarea Island (Italy) and their biotechnological potential. 827–837. Journal of Applied Microbiology, 112(6), 1102–1112. Lin, M., Yang, Y., Chen, Y., Hua, K., Lu, C., Sheu, F., ... Wu, S. (2011). A novel exopo- Gugliandolo, C., Spanò, A., Lentini, V., Arena, A., & Maugeri, T. L. (2013). Antiviral and lysaccharide from the biofilm of Thermus aquaticus YT-1 induces the immune re- immunomodulatory effects of a novel bacterial exopolysaccharide of shallow marine sponse through Toll-like receptor 2. Journal of Biological Chemistry, 286(20), vent origin. Journal of Applied Microbiology, 116(4), 1028–1034. 17736–17745. Hammer, B. K., & Bassler, B. L. (2009). Distinct sensory pathways in Vibrio cholerae El Tor Liu, P., Chen, Z., Yang, L., Li, Q., & He, N. (2017). Increasing the bioflocculant production and classical biotypes modulate cyclic dimeric GMP levels to control biofilm for- and identifying the effect of overexpressing epsB on the synthesis of polysaccharide mation. Journal of bacteriology, 191(1), 169–177. and γ-PGA in Bacillus licheniformis. Microbial Cell Factories, 16(1), 163. Han, P.-p., Sun, Y., Wu, X.-y., Yuan, Y.-j., Dai, Y.-j., & Jia, S.-r. (2014). Emulsifying, Liu, S., Chen, X., He, H., Zhang, X., Xie, B., Yu, Y., ... Zhang, Y. (2013). Structure and flocculating, and physicochemical properties of exopolysaccharide produced by cy- ecological roles of a novel exopolysaccharide from the Arctic sea ice bacterium anobacterium Nostoc flagelliforme. Applied Biochemistry and Biotechnology, 172(1), Pseudoalteromonas sp. strain SM20310. Applied and environmental microbiology, 79(1), 36–49. 224–230. Horikoshi, K. (1999). Alkaliphiles: Some applications of their products for biotechnology. Llamas, I., Amjres, H., Mata, J. A., Quesada, E., & Béjar, V. (2012). The potential bio- Microbiology and molecular biology reviews, 63(4), 735–750. technological applications of the exopolysaccharide produced by the halophilic Huang, H., Li, X., Wu, M., Wang, S., Li, G., & Ma, T. (2013). Cloning, expression and bacterium Halomonas almeriensis. Molecules, 17(6), 7103–7120. characterization of a phosphoglucomutase/phosphomannomutase from sphingan- Llamas, I., Mata, J. A., Tallon, R., Bressollier, P., Urdaci, M. C., Quesada, E., ... Béjar, V. producing Sphingomonas sanxanigenens. Biotechnology Letters, 35(8), 1265–1270. (2010). Characterization of the exopolysaccharide produced by Salipiger mucosus A3T, Huston, A. L., Methe, B., & Deming, J. W. (2004). Purification, characterization, and a halophilic species belonging to the Alphaproteobacteria, isolated on the Spanish sequencing of an extracellular cold-active aminopeptidase produced by marine psy- Mediterranean seaboard. Marine Drugs, 8(8), 2240–2251. chrophile Colwellia psychrerythraea strain 34H. Applied and environmental micro- Maleki, S., Almaas, E., Zotchev, S., Valla, S., & Ertesvåg, H. (2016). Alginate biosynthesis biology, 70(6), 3321–3328. factories in Pseudomonas fluorescens: Localization and correlation with alginate pro- Islam, S. T., & Lam, J. S. (2014). Synthesis of bacterial polysaccharides via the Wzx/Wzy- duction level. Applied and environmental microbiology, 82(4), 1227–1236. dependent pathway. Canadian Journal of Microbiology, 60(11), 697–716. Manca, M. C., Lama, L., Improta, R., Esposito, E., Gambacorta, A., & Nicolaus, B. (1996). Ito, S., Kobayashi, T., Ara, K., Ozaki, K., Kawai, S., & Hatada, Y. (1998). Alkaline de- Chemical composition of two exopolysaccharides from Bacillus thermoantarcticus. tergent enzymes from alkaliphiles: Enzymatic properties, genetics, and structures. Applied and environmental microbiology, 62(9), 3265–3269. Extremophiles, 2(3), 185–190. Martínez-Checa, F., Toledo, F. L., El Mabrouki, K., Quesada, E., & Calvo, C. (2007). Jain, R. M., Mody, K., Mishra, A., & Jha, B. (2012). Isolation and structural character- Characteristics of bioemulsifier V2-7 synthesized in culture media added of hydro- ization of biosurfactant produced by an alkaliphilic bacterium Cronobacter sakazakii carbons: Chemical composition, emulsifying activity and rheological properties. isolated from oil contaminated wastewater. Carbohydrate Polymers, 87(3), Bioresource Technology, 98(16), 3130–3135. 2320–2326. Marx, J. G., Carpenter, S. D., & Deming, J. W. (2009). Production of cryoprotectant ex- Jiang, P., Li, J., Han, F., Duan, G., Lu, X., Gu, Y., ... Yu, W. (2011). Antibiofilm activity of tracellular polysaccharide substances (EPS) by the marine psychrophilic bacterium an exopolysaccharide from marine bacterium Vibrio sp. QY101. PLOS ONE, 6(4), Colwellia psychrerythraea strain 34H under extreme conditions. Canadian Journal of e18514. Microbiology, 55(1), 63–72. Jiao, Y., Cody, G. D., Harding, A. K., Wilmes, P., Schrenk, M., Wheeler, K. E., ... Thelen, M. Mata, J. A., Béjar, V., Bressollier, P., Tallon, R., Urdaci, M., Quesada, E., ... Llamas, I. P. (2010). Characterization of extracellular polymeric substances from acidophilic (2008). Characterization of exopolysaccharides produced by three moderately halo- microbial biofilms. Applied and environmental microbiology, 76(9), 2916–2922. philic bacteria belonging to the family Alteromonadaceae. Journal of Applied Johnson, D. B. (1998). Biodiversity and ecology of acidophilic microorganisms. FEMS Microbiology, 105(2), 521–528. microbiology ecology, 27(4), 307–317. Mata, J. A., Béjar, V., Llamas, I., Arias, S., Bressollier, P., Tallon, R., ... Quesada, E. (2006). Johnson, D. B., Joulian, C., d’Hugues, P., & Hallberg, K. B. (2008). Sulfobacillus benefaciens Exopolysaccharides produced by the recently described halophilic bacteria sp. nov., an acidophilic facultative anaerobic firmicute isolated from mineral bio- Halomonas ventosae and Halomonas anticariensis. Research in Microbiology, 157(9), leaching operations. Extremophiles, 12(6), 789. 827–835. Jones, K. M. (2012). Increased production of the exopolysaccharide succinoglycan en- Maugeri, T. L., Gugliandolo, C., Caccamo, D., Panico, A., Lama, L., Gambacorta, A., ... hances Sinorhizobium meliloti 1021 symbiosis with the host plant Medicago truncatula. Nicolaus, B. (2002). A halophilic thermotolerant Bacillus isolated from a marine hot Journal of bacteriology, 194(16), 4322–4331. spring able to produce a new exopolysaccharide. Biotechnology Letters, 24(7), Junge, K., Eicken, H., Swanson, B. D., & Deming, J. W. (2006). Bacterial incorporation of 515–519. leucine into protein down to -20 degrees C with evidence for potential activity in sub- Michel, C., Beny, C., Delorme, F., Poirier, L., Spolaore, P., Morin, D., ... d’Hugues, P. eutectic saline ice formations. Cryobiology, 52(3), 417–429. (2009). New protocol for the rapid quantification of exopolysaccharides in con- Kambourova, M., Mandeva, R., Dimova, D., Poli, A., Nicolaus, B., & Tommonaro, G. tinuous culture systems of acidophilic bioleaching bacteria. Applied Microbiology and (2009). Production and characterization of a microbial , synthesized by Biotechnology, 82(2), 371–378. Geobacillus tepidamans V264 isolated from Bulgarian hot spring. Carbohydrate Moreno-Paz, M., Gómez, M. J., Arcas, A., & Parro, V. (2010). Environmental tran- Polymers, 77(2), 338–343. scriptome analysis reveals physiological differences between biofilm and planktonic Karlapudi, A. P., Kodali, V. P., Kota, K. P., Shaik, S. S., Sampath Kumar, N. S., & Dirisala, modes of life of the iron oxidizing bacteria Leptospirillum spp. in their natural mi- V. R. (2016). Deciphering the effect of novel bacterial exopolysaccharide-based na- crobial community. BMC Genomics, 11(1), 404. noparticle cream against Propionibacterium acnes.3.Biotech, 6(1), 35. Morgan, J. L. W., Strumillo, J., & Zimmer, J. (2012). Crystallographic snapshot of cel- Kim, S. J., Kim, B.-G., Park, H. J., & Yim, J. H. (2016). Cryoprotective properties and lulose synthesis and membrane translocation. Nature, 493, 181–186. preliminary characterization of exopolysaccharide (P-Arcpo 15) produced by the Moriello, V. S., Lama, L., Poli, A., Gugliandolo, C., Maugeri, T. L., Gambacorta, A., ... Arctic bacterium Pseudoalteromonas elyakovii Arcpo 15. Preparative Biochemistry and Nicolaus, B. (2003). Production of exopolysaccharides from a thermophilic micro- Biotechnology, 46(3), 261–266. organism isolated from a marine hot spring in flegrean areas. Journal of Industrial Küçükaşik, F., Kazak, H., Güney, D., Finore, I., Poli, A., Yenigün, O., ... Öner, E. T. (2011). Microbiology and Biotechnology, 30(2), 95–101. Molasses as fermentation substrate for levan production by Halomonas sp. Applied Nevot, M., Deroncele, V., Montes, M. J., & Mercade, E. (2008). Effect of incubation Microbiology and Biotechnology, 89(6), 1729–1740. temperature on growth parameters of Pseudoalteromonas antarctica NF and its pro- Le Costaouëc, T., Cérantola, S., Ropartz, D., Ratiskol, J., Sinquin, C., Colliec-Jouault, S., ... duction of extracellular polymeric substances. Journal of Applied Microbiology, 105(1), Boisset, C. (2012). Structural data on a bacterial exopolysaccharide produced by a 255–263. deep-sea Alteromonas macleodii strain. Carbohydrate Polymers, 90(1), 49–59. Nichols, C. M., Garon, S., Bowman, J. P., Raguenes, G., & Guezennec, J. (2004). Lelchat, F., Cérantola, S., Brandily, C., Colliec-Jouault, S., Baudoux, A.-C., Ojima, T., ... Production of exopolysaccharides by Antarctic marine bacterial isolates. Journal of Boisset, C. (2015). The marine bacteria Cobetia marina DSMZ 4741 synthesizes an Applied Microbiology, 96(5), 1057–1066. unexpected K-antigen-like exopolysaccharide. Carbohydrate Polymers, 124, 347–356. Nichols, C. M., Bowman, J. P., & Guezennec, J. (2005). Effects of incubation temperature Levander, F., Svensson, M., & Radstrom, P. (2002). Enhanced exopolysaccharide pro- on growth and production of exopolysaccharides by an Antarctic sea ice bacterium duction by metabolic engineering of Streptococcus thermophilus. Applied and environ- grown in batch culture. Applied and environmental microbiology, 71(7), 3519–3523. mental microbiology, 68(2), 784–790. Nichols, C. M., Guezennec, J., & Bowman, J. P. (2005). Bacterial exopolysaccharides from Li, H., Xu, H., Li, S., Xu, H., Guo, C., Ying, H., ... Ouyang, P. (2010). Strain improvement extreme marine environments with special consideration of the southern ocean, sea and metabolic flux modeling of wild-type and mutant Alcaligenes sp. NX-3 for ice, and deep-sea hydrothermal vents: A review. Marine Biotechnology, 7(4), 253–271. synthesis of exopolysaccharide welan gum. Biotechnology and Bioprocess Engineering, Nichols, C. M., Lardière, S. G., Bowman, J. P., Nichols, P. D., Gibson, J. A. E., & 15(5), 777–784. Guézennec, J. (2005). Chemical characterization of exopolysaccharides from Li, O., Qian, C., Zheng, D., Wang, P., Liu, Y., Jiang, X., ... Wu, X. (2015). Two UDP- Antarctic marine bacteria. Microbial Ecology, 49(4), 578–589. glucuronic acid decarboxylases involved in the biosynthesis of a bacterial exopoly- Nicolaus, B., Kambourova, M., & Oner, E. T. (2010). Exopolysaccharides from ex- saccharide in Paenibacillus elgii. Applied Microbiology and Biotechnology, 99(7), tremophiles: From fundamentals to biotechnology. Environmental Technology, 31(10), 3127–3139. 1145–1158. Li, P., Liu, Z., & Xu, R. (2001). Chemical characterisation of the released polysaccharide Nicolaus, B., Lama, L., Esposito, E., Manca, M. C., Improta, R., Bellitti, M. R., ... from the cyanobacterium Aphanothece halophytica GR02. Journal of Applied Phycology, Gambacorta, A. (1999). Haloarcula spp able to biosynthesize exo- and endopolymers.

24 J. Wang et al. Carbohydrate Polymers 205 (2019) 8–26

Journal of Industrial Microbiology and Biotechnology, 23(6), 489–496. (2002). Gellan gum biosynthesis in Sphingomonas paucimobilis ATCC 31461: Genes, Nicolaus, B., Lama, L., Panico, A., Moriello, V. S., Romano, I., & Gambacorta, A. (2002). enzymes and exopolysaccharide production engineering. Journal of Industrial Production and characterization of exopolysaccharides excreted by thermophilic Microbiology and Biotechnology, 29(4), 170–176. bacteria from shallow, marine hydrothermal vents of flegrean ares (Italy). Systematic Salem, D. R. (2007). Electrospinning of nanofibers and the charge injection method. In P. and Applied Microbiology, 25(3), 319–325. J. Brown, & K. Stevens (Eds.). Nanofibers and nanotechnology in textiles (pp. 3–21). Nicolaus, B., Moriello, V. S., Lama, L., Poli, A., & Gambacorta, A. (2004). Polysaccharides Woodhead Publishing. from extremophilic microorganisms. Origins of life and evolution of the biosphere, Sam, S., Kucukasik, F., Yenigun, O., Nicolaus, B., Oner, E. T., & Yukselen, M. A. (2011). 34(1), 159–169. Flocculating performances of exopolysaccharides produced by a halophilic bacterial Nicolaus, B., Panico, A., Manca, M. C., Lama, L., Gambacorta, A., Maugeri, T. L., ... strain cultivated on agro-industrial waste. Bioresource Technology, 102(2), Caccamo, D. (2000). A thermophilic Bacillus isolated from an Eolian shallow hy- 1788–1794. drothermal vent able to produce exopolysaccharides. Systematic and Applied Samain, E., Miles, M., Bozzi, L., Dubreucq, G., & Rinaudo, M. (1997). Simultaneous Microbiology, 23(3), 426–432. production of two different gel-forming exopolysaccharides by an Alteromonas strain Ollivier, B., Caumette, P., Garcia, J.-L., & Mah, R. (1994). Anaerobic bacteria from hy- originating from deep sea hydrothermal vents. Carbohydrate Polymers, 34(4), persaline environments. Microbiological reviews, 58(1), 27–38. 235–241. Panosyan, H., Di Donato, P., Poli, A., & Nicolaus, B. (2018). Production and character- Santos, C., Silva, C. J., Büttel, Z., Guimarães, R., Pereira, S. B., Tamagnini, P., ... Zille, A. ization of exopolysaccharides by Geobacillus thermodenitrificans ArzA-6 and (2014). Preparation and characterization of polysaccharides/PVA blend nanofibrous Geobacillus toebii ArzA-8 strains isolated from an Armenian geothermal spring. membranes by electrospinning method. Carbohydrate Polymers, 99, 584–592. Extremophiles, 22(5), 725–737. Sardari, R. R., Kulcinskaja, E., Ron, E. Y., Bjornsdottir, S., Friethjonsson, O. H., Perry, T. D., Klepac-Ceraj, V., Zhang, X. V., McNamara, C. J., Polz, M. F., Martin, S. T., ... Hreggviethsson, G. O., ... Karlsson, E. N. (2017). Evaluation of the production of Mitchell, R. (2005). Binding of harvested bacterial exopolymers to the surface of exopolysaccharides by two strains of the thermophilic bacterium Rhodothermus calcite. Environmental Science & Technology, 39(22), 8770–8775. marinus. Carbohydrate Polymers, 156,1–8. Poli, A., Anzelmo, G., & Nicolaus, B. (2010). Bacterial exopolysaccharides from extreme Sarilmiser, H. K., & Oner, E. T. (2014). Investigation of anti-cancer activity of linear and marine habitats: Production, characterization and biological activities. Marine Drugs, aldehyde-activated levan from Halomonas smyrnensis AAD6T. Biochemical Engineering 8(6), 1779–1802. Journal, 92,28–34. Poli, A., Kazak, H., Gürleyendağ, B., Tommonaro, G., Pieretti, G., Öner, E. T., ... Nicolaus, Sarilmiser, H. K., Ates, O., Ozdemir, G., Arga, K. Y., & Oner, E. T. (2015). Effective sti- B. (2009). High level synthesis of levan by a novel Halomonas species growing on mulating factors for microbial levan production by Halomonas smyrnensis AAD6T. defined media. Carbohydrate Polymers, 78(4), 651–657. Journal of Bioscience and Bioengineering, 119(4), 455–463. Poli, A., Moriello, V. S., Esposito, E., Lama, L., Gambacorta, A., & Nicolaus, B. (2004). Schmid, J. (2018). Recent insights in microbial exopolysaccharide biosynthesis and en- Exopolysaccharide production by a new Halomonas strain CRSS isolated from saline gineering strategies. Current opinion in biotechnology, 53, 130–136. lake Cape Russell in Antarctica growing on complex and defined media. Biotechnology Schmid, J., Sieber, V., & Rehm, B. (2015). Bacterial exopolysaccharides: Biosynthesis Letters, 26(21), 1635–1638. pathways and engineering strategies. Frontiers in Microbiology, 6, 496. Prechtl, R. M., Wefers, D., Jakob, F., & Vogel, R. F. (2018). Cold and salt stress modulate Sezer, A. D., Kazak, H., Öner, E. T., & Akbuğa, J. (2011). Levan-based nanocarrier system amount, molecular and macromolecular structure of a Lactobacillus sakei dextran. for peptide and protein drug delivery: Optimization and influence of experimental Food Hydrocolloids, 82,73–81. parameters on the nanoparticle characteristics. Carbohydrate Polymers, 84(1), Qian, C., Li, W., Li, H., Ou, D., Zhu-Ge, Y., & Liu, Y. (2018). Responses of genes for the 358–363. uptake of glycine betaine in Virgibacillus halodenitrificans PDB-F2 under NaCl stress. Sima, F., Mutlu, E. C., Eroglu, M. S., Sima, L. E., Serban, N., Ristoscu, C., ... Mihailescu, I. International Biodeterioration & Biodegradation, 132, 192–199. N. (2011). Levan nanostructured thin films by MAPLE assembling. Biomacromolecules, Qian, Y., Qi, M., Zheng, L., King, W. M., Lv, L., & Ye, F. (2016). Incorporation of rutin in 12(6), 2251–2256. electrospun pullulan/PVA nanofibers for novel UV-resistant properties. Materials, Spanò, A., & Arena, A. (2016). Bacterial exopolysaccharide of shallow marine vent origin 9(7), 504. as agent in counteracting immune disorders induced by herpes virus. Journal of Qin, G., Zhu, L., Chen, X., Wang, P. G., & Zhang, Y. (2007). Structural characterization Immunoassay and Immunochemistry, 37(3), 251–260. and ecological roles of a novel exopolysaccharide from the deep-sea psychrotolerant Spanò, A., Gugliandolo, C., Lentini, V., Maugeri, T. L., Anzelmo, G., Poli, A., ... Nicolaus, bacterium Pseudoalteromonas sp. SM9913. Microbiology, 153(Pt 5), 1566–1572. B. (2013). A novel EPS-producing strain of Bacillus licheniformis isolated from a Radchenkova, N., Boyadzhieva, I., Atanasova, N., Poli, A., Finore, I., Di Donato, P., ... shallow vent off Panarea Island (Italy). Current Microbiology, 67(1), 21–29. Kambourova, M. (2018). Extracellular polymer substance synthesized by a halophilic Spanò, A., Laganà, P., Visalli, G., Maugeri, T. L., & Gugliandolo, C. (2016). In vitro anti- bacterium Chromohalobacter canadensis 28. Applied Microbiology and Biotechnology, biofilm activity of an exopolysaccharide from the marine thermophilic Bacillus li- 102(11), 4937–4949. cheniformis T14. Current Microbiology, 72(5), 518–528. Radchenkova, N., Panchev, I., Vassilev, S., Kuncheva, M., Dobreva, S., & Kambourova, M. Stingele, F., Vincent, S. J. F., Faber, E. J., Newell, J. W., Kamerling, J. P., & Neeser, J.-R. (2015). Continuous cultivation of a thermophilic bacterium Aeribacillus pallidus 418 (1999). Introduction of the exopolysaccharide gene cluster from Streptococcus ther- for production of an exopolysaccharide applicable in cosmetic creams. Journal of mophilus Sfi6 into Lactococcus lactis MG1363: Production and characterization of an Applied Microbiology, 119(5), 1301–1309. altered polysaccharide. Molecular Microbiology, 32(6), 1287–1295. Radchenkova, N., Vassilev, S., Martinov, M., Kuncheva, M., Panchev, I., Vlaev, S., ... Subramanian, S. B., Yan, S., Tyagi, R. D., & Surampalli, R. Y. (2010). Extracellular Kambourova, M. (2014). Optimization of the aeration and agitation speed of polymeric substances (EPS) producing bacterial strains of municipal wastewater Aeribacillus palidus 418 exopolysaccharide production and the emulsifying properties sludge: Isolation, molecular identification, EPS characterization and performance for of the product. Process Biochemistry, 49(4), 576–582. sludge settling and dewatering. Water Research, 44(7), 2253–2266. Radchenkova, N., Vassilev, S., Panchev, I., Anzelmo, G., Tomova, I., Nicolaus, B., ... Sun, M.-L., Zhao, F., Shi, M., Zhang, X.-Y., Zhou, B.-C., Zhang, Y.-Z., ... Chen, X.-L. (2015). Kambourova, M. (2013). Production and properties of two novel exopolysaccharides Characterization and biotechnological potential analysis of a new exopolysaccharide synthesized by a thermophilic bacterium Aeribacillus pallidus 418. Applied from the Arctic marine bacterium Polaribacter sp. SM1127. Scientific Reports, 5, Biochemistry and Biotechnology, 171(1), 31–43. 18435. Raveendran, S., Girija, A. R., Balasubramanian, S., Ukai, T., Yoshida, Y., Maekawa, T., ... Thorne, L., Mikolajczak, M. J., Armentrout, R. W., & Pollock, T. J. (2000). Increasing the Kumar, D. S. (2014). Green approach for augmenting biocompatibility to quantum yield and viscosity of exopolysaccharides secreted by Sphingomonas by augmentation dots by extremophilic polysaccharide conjugation and nontoxic bioimaging. ACS of chromosomal genes with multiple copies of cloned biosynthetic genes. Journal of Sustainable Chemistry & Engineering, 2(7), 1551–1558. Industrial Microbiology and Biotechnology, 25(1), 49–57. Raveendran, S., Dhandayuthapani, B., Nagaoka, Y., Yoshida, Y., Maekawa, T., & Sakthi Torres-Giner, S., Ocio, M. J., & Lagaron, J. M. (2008). Development of active anti- Kumar, D. (2013). Biocompatible nanofibers based on extremophilic bacterial poly- microbial fiber-based chitosan polysaccharide nanostructures using electrospinning. saccharide, Mauran from Halomonas maura. Carbohydrate Polymers, 92(2), Engineering in Life Sciences, 8(3), 303–314. 1225–1233. Ueda, A., & Wood, T. K. (2009). Connecting quorum sensing, c-di-GMP, Pel poly- Raveendran, S., Poulose, A. C., Yoshida, Y., Maekawa, T., & Kumar, D. S. (2013). Bacterial saccharide, and biofilm formation in Pseudomonas aeruginosa through tyrosine exopolysaccharide based nanoparticles for sustained drug delivery, cancer che- phosphatase TpbA (PA3885). PLOS Pathogens, 5(6), e1000483. motherapy and bioimaging. Carbohydrate Polymers, 91(1), 22–32. Vashisth, P., Pruthi, P. A., Singh, R. P., & Pruthi, V. (2014). Process optimization for Rees, H. C., Grant, W. D., Jones, B. E., & Heaphy, S. (2004). Diversity of Kenyan soda lake fabrication of gellan based electrospun nanofibers. Carbohydrate Polymers, 109, alkaliphiles assessed by molecular methods. Extremophiles, 8(1), 63–71. 16–21. Rehm, B. H. (2010). Bacterial polymers: Biosynthesis, modifications and applications. Vu, B., Chen, M., Crawford, R. J., & Ivanova, E. P. (2009). Bacterial extracellular poly- Nature Reviews Microbiology, 8(8), 578–592. saccharides involved in biofilm formation. Molecules, 14(7), 2535–2554. Ruiz, L. M., Castro, M., Barriga, A., Jerez, C. A., & Guiliani, N. (2011). The extremophile Wang, H., Liu, Y. M., Qi, Z. M., Wang, S. Y., Liu, S. X., Li, X., ... Xia, X. C. (2013). An Acidithiobacillus ferrooxidans possesses a c‐di‐GMP signalling pathway that could play overview on natural polysaccharides with antioxidant properties. Current Medicinal a significant role during bioleaching of minerals. Letters in Applied Microbiology, 54(2), Chemistry, 20(23), 2899–2913. 133–139. Wang, L., Zhang, H., Yang, L., Liang, X., Zhang, F., & Linhardt, R. J. (2017). Structural Ruiz-Ruiz, C., Srivastava, G. K., Carranza, D., Mata, J. A., Llamas, I., Santamaría, M., ... characterization and bioactivity of exopolysaccharide synthesized by Geobacillus sp. Molina, I. J. (2011). An exopolysaccharide produced by the novel halophilic bac- TS3-9 isolated from radioactive radon hot spring. Advances in Biotechnology and terium Halomonas stenophila strain B100 selectively induces apoptosis in human T Microbiology, 4(2), 1–8. leukaemia cells. Applied Microbiology and Biotechnology, 89(2), 345–355. Welman, A. D., Maddox, I. S., & Archer, R. H. (2006). Metabolism associated with raised Rütering, M., Cress, B. F., Schilling, M., Rühmann, B., Koffas, M. A. G., Sieber, V., ... metabolic flux to sugar nucleotide precursors of exopolysaccharides in Lactobacillus Schmid, J. (2017). Tailor-made exopolysaccharides—CRISPR-Cas9 mediated genome delbrueckii subsp. bulgaricus. Journal of Industrial Microbiology and Biotechnology, editing in Paenibacillus polymyxa. Synthetic Biology, 2(1) ysx007-ysx007. 33(5), 391. Sá-Correia, I., Fialho, A. M., Videira, P., Moreira, L. M., Marques, A. R., & Albano, H. Whitney, J. C., & Howell, P. L. (2013). Synthase-dependent exopolysaccharide secretion

25 J. Wang et al. Carbohydrate Polymers 205 (2019) 8–26

in Gram-negative bacteria. Trends in Microbiology, 21(2), 63–72. Characterization of extracellular polymeric substances extracted during the bio- Xu, X., Bi, D., & Wan, M. (2016). Characterization and immunological evaluation of low- leaching of chalcopyrite concentrate. Hydrometallurgy, 100(3-4), 177–180. molecular-weight alginate derivatives. Current Topics in Medicinal Chemistry, 16(8), Zhao, S., Cao, F., Zhang, H., Zhang, L., Zhang, F., & Liang, X. (2014). Structural char- 874–887. acterization and biosorption of exopolysaccharides from Anoxybacillus sp. R4-33 Yildiz, S. Y., Anzelmo, G., Ozer, T., Radchenkova, N., Genc, S., Di Donato, P., ... isolated from radioactive radon hot spring. Applied Biochemistry and Biotechnology, Kambourova, M. (2014). Brevibacillus themoruber: A promising microbial cell factory 172(5), 2732–2746. for exopolysaccharide production. Journal of Applied Microbiology, 116(2), 314–324. Zheng, C., Li, Z., Su, J., Zhang, R., Liu, C., & Zhao, M. (2012). Characterization and Yildiz, S. Y., Radchenkova, N., Arga, K. Y., Kambourova, M., & Oner, E. T. (2015). emulsifying property of a novel bioemulsifier by Aeribacillus pallidus YM‐1. Journal of Genomic analysis of Brevibacillus thermoruber 423 reveals its biotechnological and Applied Microbiology, 113(1), 44–51. industrial potential. Applied Microbiology and Biotechnology, 99(5), 2277–2289. Zhu, P., Chen, X., Li, S., Xu, H., Dong, S., Xu, Z., ... Zhang, Y. (2014). Screening and Zeldes, B. M., Keller, M. W., Loder, A. J., Straub, C. T., Adams, M. W. W., & Kelly, R. M. characterization of Sphingomonas sp. mutant for welan gum biosynthesis at an ele- (2015). Extremely thermophilic microorganisms as metabolic engineering platforms vated temperature. Bioprocess and Biosystems Engineering, 37(9), 1849–1858. for production of fuels and industrial chemicals. Frontiers in Microbiology, 6, 1209. Zong, A., Cao, H., & Wang, F. (2012). Anticancer polysaccharides from natural resources: Zeng, W., Qiu, G., Zhou, H., Liu, X., Chen, M., Chao, W., ... Peng, J. (2010). A review of recent research. Carbohydrate Polymers, 90(4), 1395–1410.

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