Screening and Degradation Characteristics of a CP-7 Strain of Dephosphorization Bacteria

doi:10.13560/j.cnki.biotech.bull.1985.2021-1271

Abstract

Abstract:

In order to achieve the biodegradation and effective treatment of phosphorus in domestic sewage,a strain of dephosphorization bacteria was isolated from a domestic sewage treatment plant in Lanzhou to carry out enhanced biological phosphorus removal research. 1）Combined with physiological and biochemical characterization,16S rDNA sequencing analysis and phylogenetic tree construction,the results showed that the strain had a high homology with the Klebsiella oxytoca strain. 2）The bacterium was inoculated into the double screening medium at the inoculation rate of 5%（V/V）and cultured in a constant temperature oscillation incubator at 30℃ and 150 r/min. The concentration of TP was measured every 12 h. The preliminary study on the degradation characteristics of the bacterium showed that the degradation rate of TP was 58.52% within 72 h. 3）The optimal conditions of carbon source,nitrogen source and C/N were investigated. The optimal carbon source was glucose,nitrogen source was ammonium sulfate,and the optimal C/N was 5∶1. 4）By investigating the adaptability of inoculation amount,pH,temperature,initial NaCl concentration and initial phosphate concentration,it was concluded that the degradation rate of TP was the best when the inoculation amount was 8%（V/V）,pH was 9,temperature was 30℃,initial NaCl concentration was 2.5 g/L and initial phosphate concentration was 70 mg/L. The degradation rates of TP were 75.94%,90.19%,65.48%,57.02% and 52.26% respectively. At the same time,when the inoculation amount was 2%-14%（V/V）,pH was 4.0-9.0,temperature was 15-40℃,initial NaCl concentration was 0-15 g/L,initial phosphate concentration was 45-90 mg/L,the strain demonstrated varied degrees of degradation ability to TP. Correlation analysis showed that the TP degradation rate of the strain was significantly correlated with inoculation amount,pH,initial NaCl concentration and initial phosphate concentration. 5）The interaction between the strain and other strains was investigated,and the strain was mixed with a Pseudomonas aeruginosa strain. It was found that the two strains synergistically enhanced phosphorus removal. When the ratio of the two strains was 4∶1,the TP degradation rate reached 99.43%. This experiment lays a foundation for the practical application of dephosphorization bacteria in the next stage,and provides more bacterial species selection for biological enhanced phosphorus removal technology.

Key words: wastewater treatment, dephosphorization bacteria, screening and isolation, degradation characteristics

WANG Ya-jun, SI Yun-mei. Screening and Degradation Characteristics of a CP-7 Strain of Dephosphorization Bacteria[J]. Biotechnology Bulletin, 2022, 38(7): 258-268.

Figures/Tables 15

Fig.1 Colony characteristics of CP-7 strain A：Gram staining electron microscope morphology. B：Morphological characteristics of solid medium（streaked inoculation）. C：Morphological characteristics of solid medium（two-point inoculation）

Table 1 Morphological characteristics of CP-7 strain

形态特征 Morphological characteristics	CP-7
形状Shape	卵圆形Oval
菌落大小Size of the colony/mm	2-8
颜色Color	乳白色Ivory
边缘Edge	波状Undulating
光泽Luster	有Exist
质地Texture	蜡状;黏稠;湿润;易挑起Waxy,sticky,moist,easy to lift
高度Height	隆起Upheaval
透明度Pellucidity	不透明Opacity

Table 2 Physiological and biochemical characteristics of CP-7 strain

特征Characteristics	CP-7
利用葡萄糖Use of glucose	+
利用柠檬酸盐Use of citrate	+
产硫化氢Hydrogen sulfide	-
明胶液化Gelatin liquefaction	+
甲基红Methyl red	-
精氨酸双水解酶Arginine double hydrolase	-
赖氨酸脱羧酶Lysine double hydrolase	+
鸟氨酸脱羧酶Ornithine decarboxylase	-
脲酶Urease	+
V-P	+

Table 3 Identification of CP-7 strain by 16S rDNA gene sequence analysis

菌株 Strain	物种 Species	NCBI 登录号 NCBI accession No.	相似度 Percent similarity
	Klebsiella oxytoca strain JCM 1665	NR_112010.1	99.785%
CP-7	Klebsiella grimontiistrain SB73	NR_159317.1	99.427%
	Klebsiella oxytoca strain ATCC 13182	NR_118853.1	99.213%

Fig. 2 Phylogenetic tree of CP-7 strain based on 16S rDNA

Fig.3 Growth curve of CP-7 strain

Fig. 4 Effect of carbon source on the TP degradation of CP-7 strain 1：Sodium acetate. 2：Glucose. 3：Sucrose. 4：Sodium oxalate. 5：Starch

Fig. 5 Effect of nitrogen source on TP degradation of CP-7 strain 1：Sodium nitrite. 2：Ammonium sulfate. 3：Ammonium chloride. 4：Sodium nitrate. 5：Potassium nitrate

Fig. 6 Effect of C/N on the TP degradation of CP-7 strain

Fig. 7 Effect of inoculation amount on the TP degradation of CP-7 strain

Fig. 8 Effect of pH on the TP degradation of CP-7 strain

Fig. 9 Effect of temperature on the TP degradation of CP-7 strain

Fig. 10 Effect of initial NaCl concentration on the TP degradation of CP-7 strain

Fig. 11 Effect of initial phosphate concentration on the TP degradation of CP-7 strain

Fig. 12 Effect of YZ-1 strain on the TP degradation of CP-7 strain

References 29

[1]	Conley DJ, Paerl HW, Howarth RW, et al. Controlling eutrophication:nitrogen and phosphorus[J]. Science, 2009, 323(5917):1014-1015. doi: 10.1126/science.1167755 URL
[2]	Pan B, Wu J, Pan B, et al. Development of polymer-based nanosized hydrated ferric oxides(HFOs)for enhanced phosphate removal from waste effluents[J]. Water Res, 2009, 43(17):4421-4429. doi: 10.1016/j.watres.2009.06.055 URL
[3]	Zhang MY, Pan LQ, Huang F, et al. Metagenomic analysis of composition, function and cycling processes of microbial community in water, sediment and effluent of Litopenaeus vannameifarming environments under different culture modes[J]. Aquaculture, 2019, 506:280-293. doi: 10.1016/j.aquaculture.2019.03.038 URL
[4]	Yang L, Wang XH, Cui S, et al. Simultaneous removal of nitrogen and phosphorous by heterotrophic nitrification-aerobic denitrification of a metal resistant bacterium Pseudomonas putidastrain NP₅[J]. Bioresour Technol, 2019, 285:121360. doi: 10.1016/j.biortech.2019.121360 URL
[5]	Xia L, Li X, Fan W, et al. Heterotrophic nitrification and aerobic denitrification by a novel Acinetobacter sp. ND7 isolated from municipal activated sludge[J]. Bioresour Technol, 2020, 301:122749. doi: 10.1016/j.biortech.2020.122749 URL
[6]	Zhang M, Yang Q, Zhang JH, et al. Enhancement of denitrifying phosphorus removal and microbial community of long-term operation in an anaerobic anoxic oxic-biological contact oxidation system[J]. J Biosci Bioeng, 2016, 122(4):456-466. doi: 10.1016/j.jbiosc.2016.03.019 pmid: 27133708
[7]	毛世丽, 代群威, 郭军, 等. 1株高效聚磷菌的分离鉴定及固定化应用[J]. 水处理技术, 2021, 47(1):59-63.
	Mao SL, Dai QW, Guo J, et al. Isolation, identification and immobilization of a high-efficiency phosphorus accumulating bacteria[J]. Technol Water Treat, 2021, 47(1):59-63.
[8]	Tarayre C, Nguyen HT, Brognaux A, et al. Characterisation of phosphate accumulating organisms and techniques for polyphosphate detection:a review[J]. Sensors(Basel), 2016, 16(6):E797.
[9]	Seviour RJ, Mino T, Onuki M. The microbiology of biological phosphorus removal in activated sludge systems[J]. FEMS Microbiol Rev, 2003, 27(1):99-127. pmid: 12697344
[10]	Li HF, Li BZ, Wang ET, et al. Removal of low concentration of phosphorus from solution by free and immobilized cells of Pseudomonas stutzeri YG-24[J]. Desalination, 2012, 286:242-247. doi: 10.1016/j.desal.2011.11.029 URL
[11]	Liu H, Wang Q, Sun YF, et al. Isolation of a non-fermentative bacterium, Pseudomonas aeruginosa, using intracellular carbon for denitrification and phosphorus-accumulation and relevant metabolic mechanisms[J]. Bioresour Technol, 2016, 211:6-15. doi: 10.1016/j.biortech.2016.03.051 URL
[12]	亢涵, 藏春月, 焦洋, 等. 高效除磷菌P7的生长特征及除磷性能研究[J]. 沈阳建筑大学学报:自然科学版, 2017, 33(5):945-952.
	Kang H, Zang CY, Jiao Y, et al. The growth characteristics and the phosphorus removal performance of efficient phosphorus removal bacteria P7[J]. J Shenyang Jianzhu Univ:Nat Sci, 2017, 33(5):945-952.
[13]	东秀珠, 蔡妙英. 常见细菌系统鉴定手册[M]. 北京: 科学出版社, 2001.
	Dong XZ, Cai MY. Manual for systematic identification of common bacteria[M]. Beijing: Science Press, 2001.
[14]	王亚军, 蔡文娟, 耿冲冲, 等. 一株油脂降解菌的筛选及其降解条件优化[J]. 生态环境学报, 2020, 29(5):1031-1038.
	Wang YJ, Cai WJ, Geng CC, et al. Screening of a oil-degrading strain and optimization of its degradation conditions[J]. Ecol Environ Sci, 2020, 29(5):1031-1038.
[15]	蔡曼莎, 吴苇杰, 莫少庆, 等. 两种碳源对聚磷菌种群结构及除磷脱氮性能的影响[J]. 环境监测管理与技术, 2021, 33(4):65-67, 71.
	Cai MS, Wu WJ, Mo SQ, et al. Impact of two carbon sources on PAOs population structure and phosphorus and nitrogen removal performance[J]. Adm Tech Environ Monit, 2021, 33(4):65-67, 71.
[16]	庄志刚, 韩永和, 章文贤, 等. 高效聚磷菌Alcaligenes sp. -12菌株的分离鉴定及其除磷特性[J]. 环境科学学报, 2014, 34(3):678-687.
	Zhuang ZG, Han YH, Zhang WX, et al. Isolation, identification and phosphorus-removal characterization of bacteria Alcaligenes sp. strain ED-12 for phosphorus-accumulation[J]. Acta Sci Circumstantiae, 2014, 34(3):678-687.
[17]	周德庆. 微生物学教程[M]. 北京: 高等教育出版社, 1993.
	Zhou DQ. Microbiology Course[M]. Beijing: Higher Education Press, 1993.
[18]	秦华明. 高效油脂降解菌的筛选及其对油脂废水的强化处理研究[D]. 广州: 华南理工大学, 2003.
	Qin HM. Screening high effective oil degradation strain and its bioaugmentation of oil wastewater treatment[D]. Guangzhou: South China University of Technology, 2003.
[19]	崔玉波, 朱宝英. 低温下ABR酸化反应器的运行效果及微生物活性研究[J]. 环境工程学报, 2007, 1(2):49-53.
	Cui YB, Zhu BY. The performance of anaerobic baffled reactor and microorganism activity at low temperatures[J]. Chin J Environ Eng, 2007, 1(2):49-53.
[20]	杨智博, 李晓红, 张丹丹, 等. 高效油脂降解混合菌株筛选及降解条件的优化[J]. 水处理技术, 2021, 47(7):36-41.
	Yang ZB, Li XH, Zhang DD, et al. Screening of high-efficiency oil-degrading mixed strains and optimization of degradation conditions[J]. Technol Water Treat, 2021, 47(7):36-41.
[21]	吴晓娜, 王助贫, 谢恩, 等. 一株反硝化聚磷菌筛选及其接种量对脱氮除磷效应的影响[J]. 环境工程学报, 2018, 12(2):544-551.
	Wu XN, Wang ZP, Xie E, et al. Screening of one strain of denitrifying phosphorus accumulation bacteria(DPAB)and inhibitions effects of nitrogen-phosphorus removal[J]. Chin J Environ Eng, 2018, 12(2):544-551.
[22]	马瑞, 赵永鹏, 王智慧, 等. pH对水稻土全程氨氧化细菌丰度和群落结构组成影响[J]. 环境科学, 2021. DOI: https://doi.org/10.13227/j.hjkx.202107109. doi: https://doi.org/10.13227/j.hjkx.202107109
	Ma R, Zhao YP, Wang ZH, et al. Effect of pH on the abundance and community structure of comammox nitrospira in paddy soils[J]. Acta Scientiae Circumstantiae, 2021. DOI: https://doi.org/10.13227/j.hjkx.202107109. doi: https://doi.org/10.13227/j.hjkx.202107109
[23]	Coats ER, Brinkman CK, Lee S. Characterizing and contrasting the microbial ecology of laboratory and full-scale EBPR systems cultured on synthetic and real wastewaters[J]. Water Res, 2017, 108:124-136. doi: S0043-1354(16)30822-3 pmid: 27814897
[24]	Zhang T, Liu Y, Fang HHP. Effect of pH change on the performance and microbial community of enhanced biological phosphate removal process[J]. Biotechnol Bioeng, 2005, 92(2):173-182. pmid: 15962340
[25]	刘有华, 魏慧, 王倩楠, 等. 一株高效聚磷解淀粉芽孢杆菌的分离鉴定及其除磷条件优化[J]. 微生物学通报, 2021, 48(5):1473-1485.
	Liu YH, Wei H, Wang QN, et al. Isolation, identification and phosphorus removal optimization of a phosphorus-accumulating Bacillus amyloliquefaciens[J]. Microbiol China, 2021, 48(5):1473-1485.
[26]	Duan J, Fang H, Su B, et al. Characterization of a halophilic heterotrophic nitrification-aerobic denitrification bacterium and its application on treatment of saline wastewater[J]. Bioresour Technol, 2015, 179:421-428. doi: 10.1016/j.biortech.2014.12.057 URL
[27]	Akgül R, Akgül F, Kızılkaya IT. Effects of different phosphorus concentrations on growth and biochemical composition of Desmodesmus communis(E. Hegewald)E. Hegewald[J]. Prep Biochem Biotechnol, 2021, 51(7):705-713. doi: 10.1080/10826068.2020.1853156 URL
[28]	徐瑞瑞. 环境因素对群体微生物中铜绿假单胞菌与其他微生物相互作用的影响及机理探究[D]. 广州: 华南理工大学, 2020.
	Xu RR. The influence of environmental factors on the interaction between Pseudomonas aeruginosa and other microorganisms and its mechanism[D]. Guangzhou: South China University of Technology, 2020.
[29]	Abdel-Mawgoud AM, Lépine F, Déziel E. Rhamnolipids:diversity of structures, microbial origins and roles[J]. Appl Microbiol Biotechnol, 2010, 86(5):1323-1336. doi: 10.1007/s00253-010-2498-2 pmid: 20336292