北戴河海洋沉积物中L-半胱氨酸脱硫细菌的多样性及其特性
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摘要
【背景】脱硫细菌对有机硫的脱硫作用在硫的生物地球化学循环以及脱硫工业中都起着重要的作用。【目的】了解海洋沉积物中可分解有机物产生硫化氢的细菌多样性。【方法】对我国北戴河海洋沉积物中可培养的L-半胱氨酸脱硫细菌进行分离与筛选,通过对其16SrRNA基因序列测定与分析,构建系统发育树,并对其脱硫、脱氮能力进行检验。【结果】从海洋沉积物中分离得到97株细菌,从以L-半胱氨酸为硫源的培养基中筛选出62株有机脱硫专一型细菌。根据脱硫细菌的形态及其特征,从中选取12株作为典型代表做进一步分析,它们分别属于芽孢杆菌属(Bacillus)、赖氨酸芽孢杆菌属(Lysinibacillus)、动性球菌属(Planococcus)和红球菌属(Rhodococcus)。结果表明,这12株细菌均可产生半胱氨酸脱巯基酶,能够将半胱氨酸分解为丙酮酸、硫化氢和氨,即同时具备脱硫与脱氮的能力。其中有5株菌脱硫能力较强,分别属于赖氨酸芽孢杆菌属、动性球菌属和芽孢杆菌属。【结论】海洋沉积物中存在着丰富的L-半胱氨酸脱硫细菌,为进一步研究海洋中硫的生物地球化学循环提供了素材。
[Background] Organic sulfur desulfurization catalyzed by desulfurizing bacteria plays an important role in the biogeochemical cycle of sulfur and the desulfurization industry. [Objectives] In order to investigate the diversity of bacteria using the organic sulfur to produce hydrogen sulfide in marine sediments, this research focused on the isolation and characterization of desulfurizing bacteria. [Methods]The culturable desulfurizing bacteria were isolated from the marine sediments sampled from Beidaihe,which were subjected to the screening by the media containing both lead acetate and L-cysteine. Then, the16 S rRNA gene sequences of desulfurizing bacteria were retrieved against the GenBank database. All of the gene sequences with the reference sequences were used to construct the phylogenetic tree. Finally, the bacteria were tested for their capabilities for desulfurization and denitrification. [Results] 62 desulfurizing bacteria were screened out from a total of 97 bacterial strains, out of which, 12 desulfurizing bacteria were selected as the representatives to subject to phylogenetic analysis based on their 16 S rRAN gene sequences. The results indicated that all the bacterial strains belong to the genus of Bacillus,Lysinibacillus, Planococcus and Rhodococcus, respectively. Five strains with strong desulfurization ability were affiliated to three genus of Lysinibacillus, Planococcus and Bacillus. Further test showed that the desulfurizing bacteria can produce cysteine desulfhydrase to catalyze the conversion of L-cysteine to pyruvate, hydrogen sulfide and ammonia, indicating they possess both of the capabilities of desulfurization and denitrification. [Conclusion] There are abundant L-cysteine desulfurization bacteria in marine sediments, which provide materials for further study of biogeochemical cycle of sulfur in the ocean.
[Background] Organic sulfur desulfurization catalyzed by desulfurizing bacteria plays an important role in the biogeochemical cycle of sulfur and the desulfurization industry. [Objectives] In order to investigate the diversity of bacteria using the organic sulfur to produce hydrogen sulfide in marine sediments, this research focused on the isolation and characterization of desulfurizing bacteria. [Methods]The culturable desulfurizing bacteria were isolated from the marine sediments sampled from Beidaihe,which were subjected to the screening by the media containing both lead acetate and L-cysteine. Then, the16 S rRNA gene sequences of desulfurizing bacteria were retrieved against the GenBank database. All of the gene sequences with the reference sequences were used to construct the phylogenetic tree. Finally, the bacteria were tested for their capabilities for desulfurization and denitrification. [Results] 62 desulfurizing bacteria were screened out from a total of 97 bacterial strains, out of which, 12 desulfurizing bacteria were selected as the representatives to subject to phylogenetic analysis based on their 16 S rRAN gene sequences. The results indicated that all the bacterial strains belong to the genus of Bacillus,Lysinibacillus, Planococcus and Rhodococcus, respectively. Five strains with strong desulfurization ability were affiliated to three genus of Lysinibacillus, Planococcus and Bacillus. Further test showed that the desulfurizing bacteria can produce cysteine desulfhydrase to catalyze the conversion of L-cysteine to pyruvate, hydrogen sulfide and ammonia, indicating they possess both of the capabilities of desulfurization and denitrification. [Conclusion] There are abundant L-cysteine desulfurization bacteria in marine sediments, which provide materials for further study of biogeochemical cycle of sulfur in the ocean.
引文
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[14]Black KA,Dos Santos PC.Shared-intermediates in the biosynthesis of thio-cofactors:mechanisms and functions of cysteine desulfurases and sulfur acceptors[J].Biochimica et Biophysica Acta,2015,1853(6):1470-1480
[15]Oguri T,Schneider B,Reitzer L.Cysteine catabolism and cysteine desulfhydrase(CdsH/STE0458)in Salmonella enterica serovar Typhimurium[J].Journal of Bacteriology,2012,194(16):4366-4376
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[17]Fan JF,Zhang L,Ming HX,et al.PCR-RFLP analysis of bacteria16S r DNA in marine sediment of Beidaihe[J].Marine Environmental Science,2008,27(5):409-413(in Chinese)樊景凤,张兰,明红霞,等.北戴河近岸沉积物中微生物16Sr DNA的PCR-RFLP分析[J].海洋环境科学,2008,27(5):409-413
[18]Guarneros G,Ortega MV.Cysteine desulfhydrase activities of Salmonella typhimurium and Escherichia coli[J].Applied Microbiology and Biotechnology,1970,198(1):132-142
[19]Basicl A,Blomqvist M,Dahlén1 G,et al.The proteins of Fusobacterium spp.involved in hydrogen sulfide production from L-cysteine[J].BMC Microbiology,2017,17(1):61-71
[20]Stilinovi?B,Hrenovi?J.Plate method for counting proteolytic sulphide-producing bacteria[J].Acta Botanica Croatica,2009,68(1):57-66
[21]Bahugunal A,Lilyl MK,Munjal A,et al.Desulfurization of dibenzothiophene(DBT)by a novel strain Lysinibacillus sphaericus DMT-7 isolated from diesel contaminated soil[J].Journal Environmental Sciences,2011,23(6):975-982
[22]Bai HJ,Zhang ZM,Yun N,et al.Studies on removal and transformation mechanism of lead by Rhodobacter sphaeroides[J].Acta Scientiae Circumstantiae,2007,27(4):608-614(in Chinese)白红娟,张肇铭,贠妮,等.球形红细菌去除和转化铅的机理研究[J].环境科学学报,2007,27(4):608-614
[23]Bai HJ,Zhang ZM.Microbial synthesis of semiconductor lead sulfide nanoparticles using immobilized Rhodobater sphaeroides[J].Materials Letters,2009,63(9):764-766
[24]Yang Z,Lu L,Kiely CJ,et al.Biomineralized CdS quantum dot nanocrystals:optimizing synthesis conditions and improving functional properties by surface modification[J].Industrial&Engineering Chemistry Research,2016,55(43):11235-11244
[2]Hurtgen MT.The marine sulfur cycle,revisited[J].Science,2012,337(20):305-306
[3]Gupta N,Roychoudhury PK,Deb JK.Biotechnology of desulfurization of diesel:prospects and challenges[J].Applied Microbiology and Biotechnology.2005,66(4):356-366
[4]Sohrabi M,Kamyab H,Janalizadeh N,et al.Bacterial desulfurization of organic sulfur commounds exist in fossil fuels[J].Journal of Pure and Applied Microbiology,2012,6(2):717-729
[5]Ma T,Li GQ,Li J,et al.Desulfurization of dibenzothiophene by Bacillus subtilis recombinants carrying dsz ABC and dsz Dgenes[J].Biotechnology Letters,2006,28(14):1095-1100
[6]Morra MJ,Dick WA.Mechanisms of H2S production from cysteine and cysteine by microorganisms isolated from soil by selective enrichment[J].Applied and Environmental Microbiology,1991,57(5):1413-1417
[7]Ekimova GA,Fedorov DN,Tani A,et al.Distribution of1-aminocyclopropane-1-carboxylate deaminase and D-cysteine desulfhydrase genes among type species of the genus Methylobacterium[J].Antonie Van Leeuwenhoek,2018,111(3):1-12
[8]Zhang XR,Sun ZL,Wei HL,et al.Micro-biomineralization of authigenic pyrite and its implications for seafloor cold seeps[J].Marine Geology&Quaternary Geology,2017,37(2):25-32(in Chinese)张现容,孙治雷,魏合龙,等.自生换铁矿的微生物成矿机理及对冷泉泄漏的指示意义[J].海洋地质与第四纪地质,2017,37(2):25-32
[9]Qian ZG,Sun KM.Methyl sulfide in the ocean and its effect on atmospheric sulfur cycle[J].Marine Science,1984,8(2):55-59(in Chinese)钱佐国,孙昆明.海洋中的甲基硫化物及其对大气硫循环的影响[J].海洋科学,1984,8(2):55-59
[10]Mihara H,Esaki N.Bacterial cysteine desulfurases:their function and mechanisms[J].Applied Microbiology and Biotechnology,2002,60(1-2):12-23
[11]Chen L,Zhang H,Liu G,et al.First report on the bacterial diversity in the distal gut of dholes(Cuon alpinus)by using 16Sr RNA gene sequences analysis[J].Journal of Applied Genetics,2016,57(2):275-283
[12]Küster E,Williams ST.Production of hydrogen sulfide by streptomycetes and methods for its detection[J].Applied Microbiology,1964,12(1):46-52
[13]Feng YP,Qiu HN,Sun ZJ.Research progress in determination of ammonia nitrogen in water by Nessler’s reagent spectrophotome[J].Environmental Science&Technology,2016,39(S2):348-352(in Chinese)封跃鹏,邱赫男,孙自杰.纳氏试剂分光光度法测定水中氨氮研究进展[J].环境科学与技术,2016,39(S2):348-352
[14]Black KA,Dos Santos PC.Shared-intermediates in the biosynthesis of thio-cofactors:mechanisms and functions of cysteine desulfurases and sulfur acceptors[J].Biochimica et Biophysica Acta,2015,1853(6):1470-1480
[15]Oguri T,Schneider B,Reitzer L.Cysteine catabolism and cysteine desulfhydrase(CdsH/STE0458)in Salmonella enterica serovar Typhimurium[J].Journal of Bacteriology,2012,194(16):4366-4376
[16]Clarke PH.Hydrogen sulphide production by bacteria[J].Journal of General Microbiology,1953,8(3):397-407
[17]Fan JF,Zhang L,Ming HX,et al.PCR-RFLP analysis of bacteria16S r DNA in marine sediment of Beidaihe[J].Marine Environmental Science,2008,27(5):409-413(in Chinese)樊景凤,张兰,明红霞,等.北戴河近岸沉积物中微生物16Sr DNA的PCR-RFLP分析[J].海洋环境科学,2008,27(5):409-413
[18]Guarneros G,Ortega MV.Cysteine desulfhydrase activities of Salmonella typhimurium and Escherichia coli[J].Applied Microbiology and Biotechnology,1970,198(1):132-142
[19]Basicl A,Blomqvist M,Dahlén1 G,et al.The proteins of Fusobacterium spp.involved in hydrogen sulfide production from L-cysteine[J].BMC Microbiology,2017,17(1):61-71
[20]Stilinovi?B,Hrenovi?J.Plate method for counting proteolytic sulphide-producing bacteria[J].Acta Botanica Croatica,2009,68(1):57-66
[21]Bahugunal A,Lilyl MK,Munjal A,et al.Desulfurization of dibenzothiophene(DBT)by a novel strain Lysinibacillus sphaericus DMT-7 isolated from diesel contaminated soil[J].Journal Environmental Sciences,2011,23(6):975-982
[22]Bai HJ,Zhang ZM,Yun N,et al.Studies on removal and transformation mechanism of lead by Rhodobacter sphaeroides[J].Acta Scientiae Circumstantiae,2007,27(4):608-614(in Chinese)白红娟,张肇铭,贠妮,等.球形红细菌去除和转化铅的机理研究[J].环境科学学报,2007,27(4):608-614
[23]Bai HJ,Zhang ZM.Microbial synthesis of semiconductor lead sulfide nanoparticles using immobilized Rhodobater sphaeroides[J].Materials Letters,2009,63(9):764-766
[24]Yang Z,Lu L,Kiely CJ,et al.Biomineralized CdS quantum dot nanocrystals:optimizing synthesis conditions and improving functional properties by surface modification[J].Industrial&Engineering Chemistry Research,2016,55(43):11235-11244
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