Ecology and application of haloalkaliphilic anaerobic microbial communities

Haloalkaliphilic microorganisms that grow optimally at high-pH and high-salinity conditions can be found in natural environments such as soda lakes. These globally spread lakes harbour interesting anaerobic microorganisms that have the potential of being applied in existing technologies or create new opportunities. In this review, we discuss the potential application of haloalkaliphilic anaerobic microbial communities in the fermentation of lignocellulosic feedstocks material subjected to an alkaline pre-treatment, methane production and sulfur removal technology. Also, the general advantages of operation at haloalkaline conditions, such as low volatile fatty acid and sulfide toxicity, are addressed. Finally, an outlook into the main challenges like ammonia toxicity and lack of aggregation is provided.This work was performed in the TTIW- cooperation framework of Wetsus, European Centre of Excel- lence for Sustainable Water Technology (www.wetsus.nl). Wetsus is funded by the Dutch Ministry of Economic Affairs, the European Union Regional Development Fund, the Province of Fryslân, the City of Leeuwarden and the EZ/Kompas program of the“ Samenwerkingsverband Noord-Nederland”. The authors would like to thank the participants of the research theme "Sulfur", namely Paqell, for fruitful discussions and financial suppor

Propionate and butyrate dependent bacterial sulfate reduction at extremely haloalkaline conditions and description of Desulfobotulus alkaliphilus sp. nov.

Evidence on the utilization of simple fatty acids by sulfate-reducing bacteria (SRB) at extremely haloalkaline conditions are practically absent, except for a single case of syntrophy by Desulfonatronum on acetate. Our experiments with sediments from soda lakes of Kulunda Steppe (Altai, Russia) showed sulfide production with sulfate as electron acceptor and propionate and butyrate (but not acetate) as an electron donor at a pH 10–10.5 and a salinity 70–180 g l−1. With propionate as substrate, a highly enriched sulfidogenic culture was obtained in which the main component was identified as a novel representative of the family Syntrophobacteraceae. With butyrate as substrate, a pure SRB culture was isolated which oxidized butyrate and some higher fatty acids incompletely to acetate. The strain represents the first haloalkaliphilic representative of the family Desulfobacteraceae and is described as Desulfobotulus alkaliphilus sp. nov

Haloalkaliphilic spore-forming sulfidogens from soda lake sediments and description of Desulfitispora alkaliphila gen. nov., sp. nov.

An anaerobic enrichment with pyruvate as electron donor and thiosulfate at pH 10 and 0.6 M Na+ inoculated with pasteurized soda lake sediments resulted in a sulfidogenic coculture of two morphotypes of obligately anaerobic haloalkaliphilic endospore-forming clostridia, which were further isolated in pure culture. Strain AHT16 was a thin long rod able to ferment sugars and pyruvate and to respire H2, formate and pyruvate using thiosulfate and fumarate as electron acceptors and growing optimally at pH 9.5. Thiosulfate was reduced incompletely to sulfide and sulfite. The strain was closely related (99% sequence similarity) to a peptolytic alkaliphilic clostridium Natronincola peptidovorans. Strain AHT17 was a short rod with a restricted respiratory metabolism, growing with pyruvate and lactate as electron donor and sulfite, thiosulfate and elemental sulfur as electron acceptors with a pH optimum 9.5. Thiosulfate was reduced completely via sulfite to sulfide. The ability of AHT17 to use sulfite explained the stability of the original coculture of the two clostridia—one member forming sulfite from thiosulfate and another consuming it. Strain AHT17 formed an independent deep phylogenetic lineage within the Clostridiales and is proposed as a new genus and species Desulfitisporum alkaliphilum gen. nov., sp. nov. (=DSM 22410T = UNIQEM U794T)

Anaerobic utilization of pectinous substrates at extremely haloalkaline conditions by Natranaerovirga pectinivora gen. nov., sp. nov., and Natranaerovirga hydrolytica sp. nov., isolated from hypersaline soda lakes

Anaerobic enrichments at pH 10, with pectin and polygalacturonates as substrates and inoculated with samples of sediments of hypersaline soda lakes from the Kulunda Steppe (Altai, Russia) demonstrated the potential for microbial pectin degradation up to soda-saturating conditions. The enrichments resulted in the isolation of six strains of obligately anaerobic fermentative bacteria, which represented a novel deep lineage within the order Clostridiales loosely associated with the family Lachnospiraceae. The isolates were rod-shaped and formed terminal round endospores. One of the striking features of the novel group is a very narrow substrate spectrum for growth, restricted to galacturonic acid and its polymers (e.g. pectin). Acetate and formate were the final fermentation products. Growth was possible in a pH range from 8 to 10.5, with an optimum at pH 9.5–10, and in a salinity range from 0.2 to 3.5 M Na+. On the basis of unique phenotypic properties and distinct phylogeny, the pectinolytic isolates are proposed to be assigned to a new genus Natranaerovirga with two species N. hydrolytica (APP2T=DSM24176T=UNIQEM U806T) and N. pectinivora (AP3T=DSM24629T=UNIQEM U805T)

Desulfonatronovibrio halophilus sp. nov., a novel moderately halophilic sulfate-reducing bacterium from hypersaline chloride–sulfate lakes in Central Asia

Four strains of lithotrophic sulfate-reducing bacteria (SRB) have been enriched and isolated from anoxic sediments of hypersaline chloride–sulfate lakes in the Kulunda Steppe (Altai, Russia) at 2 M NaCl and pH 7.5. According to the 16S rRNA gene sequence analysis, the isolates were closely related to each other and belonged to the genus Desulfonatronovibrio, which, so far, included only obligately alkaliphilic members found exclusively in soda lakes. The isolates utilized formate, H2 and pyruvate as electron donors and sulfate, sulfite and thiosulfate as electron acceptors. In contrast to the described species of the genus Desulfonatronovibrio, the salt lake isolates could only tolerate high pH (up to pH 9.4), while they grow optimally at a neutral pH. They belonged to the moderate halophiles growing between 0.2 and 2 M NaCl with an optimum at 0.5 M. On the basis of their distinct phenotype and phylogeny, the described halophilic SRB are proposed to form a novel species within the genus Desulfonatronovibrio, D. halophilus (type strain HTR1T = DSM24312T = UNIQEM U802T)

Desulfurispira natronophila gen. nov. sp. nov.: an obligately anaerobic dissimilatory sulfur-reducing bacterium from soda lakes

Anaerobic enrichment cultures with elemental sulfur as electron acceptor and either acetate or propionate as electron donor and carbon source at pH 10 and moderate salinity inoculated with sediments from soda lakes in Kulunda Steppe (Altai, Russia) resulted in the isolation of two novel members of the bacterial phylum Chrysiogenetes. The isolates, AHT11 and AHT19, represent the first specialized obligate anaerobic dissimilatory sulfur respirers from soda lakes. They use either elemental sulfur/polysulfide or arsenate as electron acceptor and a few simple organic compounds as electron donor and carbon source. Elemental sulfur is reduced to sulfide through intermediate polysulfide, while arsenate is reduced to arsenite. The bacteria belong to the obligate haloalkaliphiles, with a pH growth optimum from 10 to 10.2 and a salt range from 0.2 to 3.0 M Na+ (optimum 0.4–0.6 M). According to the phylogenetic analysis, the two strains were close to each other, but distinct from the nearest relative, the haloalkaliphilic sulfur-reducing bacterium Desulfurispirillum alkaliphilum, which was isolated from a bioreactor. On the basis of distinct phenotype and phylogeny, the soda lake isolates are proposed as a new genus and species, Desulfurispira natronophila (type strain AHT11T = DSM22071T = UNIQEM U758T)

Desulfuribacillus alkaliarsenatis gen. nov. sp. nov., a deep-lineage, obligately anaerobic, dissimilatory sulfur and arsenate-reducing, haloalkaliphilic representative of the order Bacillales from soda lakes

An anaerobic enrichment culture inoculated with a sample of sediments from soda lakes of the Kulunda Steppe with elemental sulfur as electron acceptor and formate as electron donor at pH 10 and moderate salinity inoculated with sediments from soda lakes in Kulunda Steppe (Altai, Russia) resulted in the domination of a Gram-positive, spore-forming bacterium strain AHT28. The isolate is an obligate anaerobe capable of respiratory growth using elemental sulfur, thiosulfate (incomplete reduction) and arsenate as electron acceptor with H2, formate, pyruvate and lactate as electron donor. Growth was possible within a pH range from 9 to 10.5 (optimum at pH 10) and a salt concentration at pH 10 from 0.2 to 2 M total Na+ (optimum at 0.6 M). According to the phylogenetic analysis, strain AHT28 represents a deep independent lineage within the order Bacillales with a maximum of 90 % 16S rRNA gene similarity to its closest cultured representatives. On the basis of its distinct phenotype and phylogeny, the novel haloalkaliphilic anaerobe is suggested as a new genus and species, Desulfuribacillus alkaliarsenatis (type strain AHT28T = DSM24608T = UNIQEM U855T)

Citric acid wastewater as electron donor for biological sulfate reduction

Citrate-containing wastewater is used as electron donor for sulfate reduction in a biological treatment plant for the removal of sulfate. The pathway of citrate conversion coupled to sulfate reduction and the microorganisms involved were investigated. Citrate was not a direct electron donor for the sulfate-reducing bacteria. Instead, citrate was fermented to mainly acetate and formate. These fermentation products served as electron donors for the sulfate-reducing bacteria. Sulfate reduction activities of the reactor biomass with acetate and formate were sufficiently high to explain the sulfate reduction rates that are required for the process. Two citrate-fermenting bacteria were isolated. Strain R210 was closest related to Trichococcus pasteurii (99.5% ribosomal RNA (rRNA) gene sequence similarity). The closest relative of strain S101 was Veillonella montepellierensis with an rRNA gene sequence similarity of 96.7%. Both strains had a complementary substrate range