Isolation and characterization of two novel alkalitolerant sulfidogens from a Thiopaq bioreactor, Desulfonatronum alkalitolerans sp. nov., and Sulfurospirillum alkalitolerans sp. nov

Two obligately anaerobic sulfidogenic bacterial strains were isolated from the full-scale Thiopaq bioreactor in Lelystad (The Netherlands) removing H2S from biogas under oxygen-limiting and moderately haloalkaline conditions. Strain HSRB-L represents a dominant culturable sulfate-reducing bacterium in the reactor. It utilizes formate, H2 (with acetate as C-source) and lactate as e-donors, and sulfate, thiosulfate and sulfite as e-acceptors. It is haloalkalitolerant, with a pH range for lithotrophic growth from 7.5 to 9.7 (optimum at 8.5–9) and a salt range from 0.1 to 1.75 M total Na+ (optimum at 0.6 M). The strain is a member of the genus Desulfonatronum and is proposed as a novel species D. alkalitolerans. The second strain, strain HTRB-L1, represents a dominant thiosulfate/sulfur reducer in the reactor. It is an obligate anaerobe utilizing formate and H2 (with acetate as C-source), lactate, pyruvate and fumarate as e-donors, and thiosulfate (incomplete reduction), sulfur, arsenate and fumarate as e-acceptors. With lactate as e-donor it also grows as an ammonifyer in the presence of nitrate and nitrite. HTRB-L1 is haloalkalitolerant, with a pH range for lithotrophic growth from 7.1 to 9.7 (optimum at 8.5) and a salt range from 0.6 to 1.5 M total Na+ (optimum at 0.6 M). Phylogenetic analysis showed that strain HTRB-L1 is a novel species within the genus Sulfurospirillum (Epsilonproteobacteria) for which a name Sulfurospirillum alkalitolerans is proposed.

[1]  F. Rainey,et al.  Desulfonatronum lacustre gen. nov., sp. nov. : A new alkaliphilic sulfate-reducing bacterium utilizing ethanol , 1998 .

[2]  C. Buisman,et al.  Application of bacteria involved in the biological sulfur cycle for paper mill effluent purification. , 2009, The Science of the total environment.

[3]  W. D. de Vos,et al.  Description of Sulfurospirillum halorespirans sp. nov., an anaerobic, tetrachloroethene-respiring bacterium, and transfer of Dehalospirillum multivorans to the genus Sulfurospirillum as Sulfurospirillum multivorans comb. nov. , 2003, International journal of systematic and evolutionary microbiology.

[4]  H. G. Trüper,et al.  Sulphur metabolism in Thiorhodaceae I. Quantitative measurements on growing cells ofChromatium okenii , 2005, Antonie van Leeuwenhoek.

[5]  P. Doty,et al.  Determination of the base composition of deoxyribonucleic acid from its thermal denaturation temperature. , 1962, Journal of molecular biology.

[6]  B. Berks,et al.  Specificity of respiratory pathways involved in the reduction of sulfur compounds by Salmonella enterica. , 2002, Microbiology.

[7]  Yves Van de Peer,et al.  TREECON for Windows: a software package for the construction and drawing of evolutionary trees for the Microsoft Windows environment , 1994, Comput. Appl. Biosci..

[8]  Kazuya Watanabe,et al.  Sulfurospirillum cavolei sp. nov., a facultatively anaerobic sulfur-reducing bacterium isolated from an underground crude oil storage cavity. , 2007, International journal of systematic and evolutionary microbiology.

[9]  G. Muyzer,et al.  Denaturing gradient gel electrophoresis in marine microbial ecology , 2001 .

[10]  P. Kroneck,et al.  Comparative systematic study on “Spirillum” 5175, Campylobacter and Wolinella species , 1992, Archives of Microbiology.

[11]  R. Mackie,et al.  Denaturing gradient gel electrophoresis , 2005 .

[12]  W. John Über das Vitamin E , 1939 .

[13]  F. Rainey,et al.  Desulfonatronovibrio hydrogenovorans gen. nov., sp. nov., an alkaliphilic, sulfate-reducing bacterium. , 1997, International journal of systematic bacteriology.

[14]  O. H. Lowry,et al.  Protein measurement with the Folin phenol reagent. , 1951, The Journal of biological chemistry.

[15]  G. Muyzer,et al.  Thioalkalivibrio sulfidiphilus sp. nov., a haloalkaliphilic, sulfur-oxidizing gammaproteobacterium from alkaline habitats. , 2012, International journal of systematic and evolutionary microbiology.

[16]  J. G. Kuenen,et al.  The Microbial Sulfur Cycle at Extremely Haloalkaline Conditions of Soda Lakes , 2011, Front. Microbio..

[17]  M. W. Weatherburn Phenol-hypochlorite reaction for determination of ammonia , 1967 .

[18]  P. Kroneck,et al.  Microbial sulfite respiration. , 2013, Advances in microbial physiology.

[19]  D. Lovley,et al.  Sulfurospirillum barnesii sp. nov. and Sulfurospirillum arsenophilum sp. nov., new members of the Sulfurospirillum clade of the epsilon Proteobacteria. , 1999, International journal of systematic bacteriology.

[20]  N. Pfennig,et al.  Über das Vitamin B12-Bedürfnis phototropher Schwefelbakterien , 1966, Archiv für Mikrobiologie.

[21]  G. Muyzer,et al.  Culturable diversity of lithotrophic haloalkaliphilic sulfate-reducing bacteria in soda lakes and the description of Desulfonatronumthioautotrophicum sp. nov., Desulfonatronum thiosulfatophilum sp. nov., Desulfonatronovibrio thiodismutans sp. nov., and Desulfonatronovibrio magnus sp. nov. , 2011, Extremophiles.

[22]  Damien Marsic,et al.  Desulfonatronum thiodismutans sp. nov., a novel alkaliphilic, sulfate-reducing bacterium capable of lithoautotrophic growth. , 2003, International journal of systematic and evolutionary microbiology.

[23]  B. Tindall,et al.  Desulfurispirillum alkaliphilum gen. nov. sp. nov., a novel obligately anaerobic sulfur- and dissimilatory nitrate-reducing bacterium from a full-scale sulfide-removing bioreactor , 2007, Extremophiles.

[24]  Gatze Lettinga,et al.  Removal of hydrogen sulphide from wastewater and waste gases by biological conversion to elemental sulphur colloidal and interfacial aspects of biologically produced sulphur particles , 1999 .

[25]  R. Mackie,et al.  4.1. Denaturing gradient gel electrophoresis , 2005 .

[26]  C. Plugge Anoxic media design, preparation, and considerations. , 2005, Methods in enzymology.

[27]  J. Marmur A procedure for the isolation of deoxyribonucleic acid from micro-organisms , 1961 .

[28]  J. Kuever,et al.  Desulfonatronum cooperativum sp. nov., a novel hydrogenotrophic, alkaliphilic, sulfate-reducing bacterium, from a syntrophic culture growing on acetate. , 2005, International journal of systematic and evolutionary microbiology.