Microlunatus Phosphovorus Research Articles

Metabolisms of Microlunatus phosphovorus NM-1 Using Glucose, Glutamate, and Aspartate as Carbon Sources for Enhanced Biological Phosphorus Removal

Metabolisms of <i>Microlunatus phosphovorus</i> NM-1 Using Glucose, Glutamate, and Aspartate as Carbon Sources for Enhanced Biological Phosphorus Removal

Carbon uptake bioenergetics of PAOs and GAOs in full-scale enhanced biological phosphorus removal systems

This work analyzed, for the first time, the bioenergetics of PAOs and GAOs in full-scale wastewater treatment plants (WWTPs) for the uptake of different carbon sources. Fifteen samples were collected from five full-scale WWTPs. Predominance of different PAOs, i.e., Ca. Accumulibacter (0.00–0.49%), Tetrasphaera (0.37–3.94%), Microlunatus phosphovorus (0.01–0.18%), etc., and GAOs, i.e., Ca. Competibacter (0.08–5.39%), Defluviicoccus (0.05–5.34%), Micropruina (0.17–1.87%), etc., were shown by 16S rRNA gene amplicon sequencing. Despite the distinct PAO/GAO community compositions in different samples, proton motive force (PMF) was found as the key driving force (up to 90.1%) for the uptake of volatile fatty acids (VFAs, acetate and propionate) and amino acids (glutamate and aspartate) by both GAOs and PAOs at the community level, contrasting the previous understanding that Defluviicoccus have a low demand of PMF for acetate uptake. For the uptake of acetate or propionate, PAOs rarely activated F1, F0- ATPase (< 11.7%) or fumarate reductase (< 5.3%) for PMF generation; whereas, intensive involvements of these two pathways (up to 49.2% and 61.0%, respectively) were observed for GAOs, highlighting a major and community-level difference in their VFA uptake biogenetics in full-scale systems. However, different from VFAs, the uptake of glutamate and aspartate by both PAOs and GAOs commonly involved fumarate reductase and F1, F0-ATPase activities. Apart from these major and community-level differences, high level fine-scale micro-diversity in carbon uptake bioenergetics was observed within PAO and GAO lineages, probably resulting from their versatilities in employing different pathways for reducing power generation. Ca. Accumulibacter and Halomonas seemed to show higher dependency on the reverse operation of F1, F0-ATPase than other PAOs, likely due to the low involvement of glyoxylate shunt pathway. Unlike Tetrasphaera, but similar to Ca. Accumulibacter, Microlunatus phosphovorus took up glutamate and aspartate via the proton/glutamate-aspartate symporter driven by PMF. This feature was testified using a pure culture of Microlunatus phosphovorus stain NM-1. The major difference between PAOs and GAOs highlights the potential to selectively suppress GAOs for community regulation in EBPR systems. The finer-scale carbon uptake bioenergetics of PAOs or GAOs from different lineages benefits in understanding their interactions in community assembly in complex environment.

Classroom microbiome, functional pathways and sick-building syndrome (SBS) in urban and rural schools - Potential roles of indoor microbial amino acids and vitamin metabolites

Sick building symptoms (SBS) are defined as non-specific symptoms related to indoor exposures, including mucosal symptoms in eye, nose, throat, and skin, and general symptoms as headache and tiredness. Indoor microbial composition is associated with SBS symptoms, but the impact of microbial functional genes and potential metabolic products has not been characterized. We conducted a shotgun microbial metagenomic sequencing for vacuum dust collected in urban and rural schools in Shanxi province, China. SBS symptoms in students were surveyed, and microbial taxa and functional pathways related to the symptoms were identified using a multi-level linear regression model. SBS symptoms were common in students, and the prevalence of ocular and throat symptoms, headache, and tiredness was higher in urban than in rural areas (p < 0.05). A significant higher microbial α-diversity was found in rural areas than in urban areas (Chao1, p = 0.001; ACE, p = 0.002). Also, significant variation in microbial taxonomic and functional composition (β-diversity) was observed between urban and rural areas (p < 0.005). Five potential risk Actinobacteria species were associated with SBS symptoms (p < 0.01); students in the classrooms with a higher abundance of an unclassified Geodermatophilaceae, Geodermatophilus, Fridmanniella luteola, Microlunatus phosphovorus and Mycetocola reported more nasal and throat symptoms and tiredness. Students with a higher abundance of an unclassified flavobacteriaceae reported fewer throat symptoms and tiredness. The abundance of microbial metabolic pathways related to the synthesis of B vitamins (biotin and folate), gamma-aminobutyric acid (GABA), short-chain fatty acids (SCFAs), and peptidoglycan and were protectively (negatively) associated with SBS symptoms (FDR < 0.05). The result is consistent with human microbiota studies, which reported that these microbial products are extensively involved in immunological processes and anti-inflammatory effects. This is the first study to report the functional potential of the indoor microbiome and the occurrence of SBS, providing new insights into the potential etiologic mechanisms in chronic inflammatory diseases.

The PolS-PolR Two-Component System Regulates Genes Involved in Poly-P Metabolism and Phosphate Transport in Microlunatus phosphovorus.

Microlunatus phosphovorus NM-1 is a polyphosphate (poly-P)-accumulating bacterium that accumulates poly-P under aerobic conditions and degrades poly-P under anaerobic conditions. In this study, the two-component system (TCS) PolS-PolR was identified in NM-1, and the response regulator PolR was found to directly bind to the promoters of genes related to phosphate transport (MLP_RS00235, MLP_RS23035, and MLP_RS24590); poly-P catabolism (MLP_RS12905) and poly-P synthesis (MLP_RS23025). RT-qPCR assays showed that ppgk (MLP_RS12905), ppk (MLP_RS23025), pstS (MLP_RS23035), and pit (MLP_RS24590) were down-regulated during the aerobic-anaerobic shift. The sequence GTTCACnnnnnGTTCaC was identified as a recognition sequence for PolR by MEME analysis and DNase I footprinting. EMSAs and ChIP-qPCR assays indicated that PolR binds to the promoters of pit (MLP_RS00235), ppgk (MLP_RS12905), ppk (MLP_RS23025), pstS (MLP_RS23035) and pit (MLP_RS24590), and ChIP-qPCR further suggested that the binding affinity of PolR was lower under anaerobic conditions than under aerobic conditions in vivo. These findings indicate that the PolS-PolR TCS in M. phosphovorus may be involved in the regulation of poly-P metabolism in response to levels of dissolved oxygen in the environment, and our results provide insights into new approaches for understanding the mechanisms of phosphorus accumulation and release.

Polyphosphate metabolic gene expression analyses reveal mechanisms of phosphorus accumulation and release in Microlunatus phosphovorus strain JN459.

The ability of Microlunatus phosphovorus to accumulate large amounts of polyphosphate (Poly-P) plays an important role in removing soluble phosphorus from wastewater. Strain JN459, isolated from a sewage system, was previously demonstrated to be Microlunatus phosphovorus. In this study, we analyzed the phosphorus-accumulating and phosphorus-releasing characteristics of strain JN459. Our analyses indicate that strain JN459 accumulates Poly-P under aerobic conditions but releases phosphorus under anaerobic conditions. To determine the mechanisms underlying Poly-P metabolism in strain JN459, we compared transcriptional profiles under aerobic and anaerobic conditions. Significant differences were detected in the expression levels of genes associated with Poly-P metabolism between aerobic and anaerobic conditions, including ppk (MLP_47700, MLP_50300 and MLP_05750), ppgk (MLP_05430 and MLP_26610), ppx (MLP_44770), pap (MLP_23310) and ppnk (MLP_17420). The high expression of polyphosphate glucokinase (MLP_05430) and polyphosphate/ATP-dependent NAD kinase (MLP_17420) indicated that both of them might be responsible for utilizing Poly-P as the energy resource for growth under anaerobic conditions. These findings enhance our understanding of phosphate metabolism in a major bacterial species involved in wastewater phosphorus reduction.

Metagenomic characterization of ‘Candidatus Defluviicoccus tetraformis strain TFO71’, a tetrad‐forming organism, predominant in an anaerobic–aerobic membrane bioreactor with deteriorated biological phosphorus removal

In an acetate-fed anaerobic-aerobic membrane bioreactor with deteriorated enhanced biological phosphorus removal (EBPR), Defluviicoccus-related tetrad-forming organisms (DTFO) were observed to predominate in the microbial community. Using metagenomics, a partial genome of the predominant DTFO, 'Candidatus Defluviicoccus tetraformis strain TFO71', was successfully constructed and characterized. Examining the genome confirmed the presence of genes related to the synthesis and degradation of glycogen and polyhydroxyalkanoate (PHA), which function as energy and carbon storage compounds. TFO71 and 'Candidatus Accumulibacter phosphatis' (CAP) UW-1 and CAP UW-2, representative polyphosphate-accumulating organisms (PAO), have PHA metabolism-related genes with high homology, but TFO71 has unique genes for PHA synthesis, gene regulation and granule management. We further discovered genes encoding DTFO polyphosphate (polyP) synthesis, suggesting that TFO71 may synthesize polyP under untested conditions. However, TFO71 may not activate these genes under EBPR conditions because the retrieved genome does not contain all inorganic phosphate transporters that are characteristic of PAOs (CAP UW-1, CAP UW-2, Microlunatus phosphovorus NM-1 and Tetrasphaera species). As a first step in characterizing EBPR-associated DTFO metabolism, this study identifies important differences between DTFO and PAO that may contribute to EBPR community competition and deterioration.

Deciphering the Genome of Polyphosphate Accumulating Actinobacterium Microlunatus phosphovorus

Polyphosphate accumulating organisms (PAOs) belong mostly to Proteobacteria and Actinobacteria and are quite divergent. Under aerobic conditions, they accumulate intracellular polyphosphate (polyP), while they typically synthesize polyhydroxyalkanoates (PHAs) under anaerobic conditions. Many ecological, physiological, and genomic analyses have been performed with proteobacterial PAOs, but few with actinobacterial PAOs. In this study, the whole genome sequence of an actinobacterial PAO, Microlunatus phosphovorus NM-1T (NBRC 101784T), was determined. The number of genes for polyP metabolism was greater in M. phosphovorus than in other actinobacteria; it possesses genes for four polyP kinases (ppks), two polyP-dependent glucokinases (ppgks), and three phosphate transporters (pits). In contrast, it harbours only a single ppx gene for exopolyphosphatase, although two copies of ppx are generally present in other actinobacteria. Furthermore, M. phosphovorus lacks the phaABC genes for PHA synthesis and the actP gene encoding an acetate/H+ symporter, both of which play crucial roles in anaerobic PHA accumulation in proteobacterial PAOs. Thus, while the general features of M. phosphovorus regarding aerobic polyP accumulation are similar to those of proteobacterial PAOs, its anaerobic polyP use and PHA synthesis appear to be different.

Microbial selection on enhanced biological phosphorus removal systems fed exclusively with glucose

The microbial selection on an enhanced biological phosphorus removal (EBPR) system was investigated in a laboratory-scale sequencing batch reactor fed exclusively with glucose as the carbon source. Fluorescence In Situ Hybridization analysis was performed to target two polyphosphate accumulating organisms (PAOs) (i.e., Candidatus Accumulibacter phosphatis and Microlunatus phosphovorus) and two glycogen accumulating organisms (GAOs) (i.e., Candidatus Competibacter phosphatis and Micropruina glycogenica). The results show that glucose might not select for Candidatus Accumulibacter phosphatis. However, Microlunatus phosphovorus, Candidatus Competibacter phosphatis, and Micropruina glycogenica might be selected. The highest percent relative abundance (% RA) of Candidatus Accumulibacter phosphatis was about 42%; this occurred at the beginning of the experimental period when phosphorus removal was efficient. However, the % RA of these bacteria decreased, reaching below 4% at the end of the run. The maximum % RA of Microlunatus phosphovorus, Candidatus Competibacter phosphatis, and Micropruina glycogenica was about 21, 37, 17%, respectively. It appears that a higher glucose concentration might be detrimental for Microlunatus phosphovorus and Micropruina glycogenica. Results also indicate a dominance of GAOs over PAOs when EBPR systems are fed with glucose. It is possible that the GAOs outcompete the PAOs at low pH values; it has been reported that at low pH, GAOs use glycogen as the energy source to uptake glucose. As a result, P-removal deteriorated. Therefore, glucose is not a strong candidate as a carbon source to supplement EBPR systems that do not contain sufficient volatile fatty acids.

Development of Quantitative PCR method for Microlunatus phosphovorus and Its Application to Activated Sludge Sample

Development of Quantitative PCR method for Microlunatus phosphovorus and Its Application to Activated Sludge Sample

Microlunatus parietis sp. nov., isolated from an indoor wall

A Gram-positive, coccoid, non-endospore-forming actinobacterium (strain 12-Be-011(T)) was isolated from indoor wall material. Based on 16S rRNA gene sequence comparisons, strain 12-Be-011(T) was clearly shown to belong to the genus Microlunatus and was most closely related to Microlunatus panaciterrae Gsoil 954(T) (95.7 %), Microlunatus soli CC-12602(T) (94.9 %), Microlunatus ginsengisoli Gsoil 633(T) (94.8 %), Microlunatus aurantiacus YIM 45721(T) (95.5 %) and Microlunatus phosphovorus DSM 10555(T) (94.7 %). The cell-wall peptidoglycan contained LL-diaminopimelic acid as the diagnostic diamino acid. Mycolic acids were absent. The major menaquinone was MK-9(H₄). The polar lipid profile consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, two unknown phospholipids and one unknown glycolipid. The major fatty acids of iso-C₁₅:₀, anteiso-C₁₅:₀ and iso-C₁₆:₀ supported the affiliation of strain 12-Be-011(T) to the genus Microlunatus. Physiological and biochemical test results allowed a clear phenotypic differentiation of strain 12-Be-011(T) from all other species of the genus Microlunatus. Hence, strain 12-Be-011(T) can be regarded as a representative of a novel species, for which the name Microlunatus parietis sp. nov. is proposed, with the type strain 12-Be-011(T) (=DSM 22083(T)=CCM 7636(T)).

Microlunatus panaciterrae sp. nov., a -glucosidase-producing bacterium isolated from soil in a ginseng field

A novel bacterial strain, designated Gsoil 954(T), showing beta-glucosidase activity was isolated from a soil sample from a ginseng field in Pocheon Province, South Korea. The isolate was a Gram-positive, aerobic, motile, coccus-shaped, non-endospore-forming bacterium. Comparative analysis of 16S rRNA gene sequences revealed that strain Gsoil 954(T) belonged to the family Propionibacteriaceae, the highest levels of sequence similarity being found with the type strains of Microlunatus ginsengisoli (96.4 %) and Microlunatus phosphovorus (96.2 %). The strain showed <95.9 % similarity with respect to other species within the family Propionibacteriaceae. In addition, the presence of ll-2,6-diaminopimelic acid in the cell-wall peptidoglycan, MK-9(H(4)) as the major menaquinone and anteiso-C(15 : 0) and iso-C(15 : 0) as the major cellular fatty acids suggested its affiliation with the genus Microlunatus. The G+C content of the genomic DNA was 65.1 mol%. On the basis of its phenotypic characteristics and phylogenetic distinctiveness, strain Gsoil 954(T) represents a novel species within the genus Microlunatus, for which the name Microlunatus panaciterrae sp. nov. is proposed. The type strain is Gsoil 954(T) (=KCTC 13058(T)=DSM 18662(T)).

Microlunatus aurantiacus sp. nov., a novel actinobacterium isolated from a rhizosphere soil sample

A Gram-positive, aerobic, non-motile, mesophilic strain, designated YIM 45721T, was isolated from a rhizosphere soil sample in Yunnan Province, China. On the basis of 16S rRNA gene sequence similarity analysis, strain YIM 45721T was assigned to the genus Microlunatus and was most closely related to the type strain of Microlunatus phosphovorus (98.2 % 16S rRNA gene sequence similarity). Chemotaxonomic data, including cell-wall peptidoglycan structure, menaquinones and fatty acid profile, supported affiliation of strain YIM 45721T to the genus Microlunatus. Results of DNA-DNA hybridization and physiological and biochemical tests enabled strain YIM 45721T to be differentiated genotypically and phenotypically from M. phosphovorus. Therefore, strain YIM 45721T represents a novel species of the genus Microlunatus, for which the name Microlunatus aurantiacus sp. nov. is proposed. The type strain is YIM 45721T (=CCTCC AB 206067T=DSM 18424T).

Microlunatus ginsengisoli sp. nov., isolated from soil of a ginseng field

A Gram-positive, aerobic, coccus-shaped, non-endospore-forming bacterium (Gsoil 633(T)) was isolated from soil from a ginseng field in Pocheon province in South Korea. The novel isolate was characterized in order to determine its taxonomic position. On the basis of 16S rRNA gene sequence similarities, strain Gsoil 633(T) was shown to belong to the family Propionibacteriaceae. The closest phylogenetic relative was Microlunatus phosphovorus DSM 19555(T), with 96.1 % sequence similarity; the sequence similarity to other members of the family was less than 95.4 %. The isolate was characterized chemotaxonomically as having ll-2,6-diaminopimelic acid in the cell-wall peptidoglycan, MK-9(H(4)) as the predominant menaquinone and anteiso-C(15 : 0), iso-C(15 : 0) and iso-C(16 : 0) as the major fatty acids. The G+C content of the genomic DNA was 69.8 mol%. The morphological and chemotaxonomic properties of the isolate were consistent with those of M. phosphovorus, but the results of physiological and biochemical tests allowed the phenotypic differentiation of strain Gsoil 633(T) from this species. Therefore, strain Gsoil 633(T) represents a novel species, for which the name Microlunatus ginsengisoli sp. nov. is proposed. The type strain is Gsoil 633(T) (=KCTC 13940(T)=DSM 17942(T)).

Isolation, characterization of bacteriophages specific to Microlunatus phosphovorus and their application for rapid host detection

To isolate and characterize lytic-bacteriophages specific to Microlunatus phosphovorus, and prepare fluorescently labelled phages (FLPs) for the rapid detection of the host bacterium in activated sludge. Isolation of bacteriophages lytic to M. phosphovorus was attempted by applying supernatants of activated sludge processes on the lawn of M. phosphovorus JCM9379 for plaque formation. Thirteen bacteriophage isolates were obtained. The restriction fragment length polymorphism analysis distinguished them into two different bacteriophages designated as phiMP1 and phiMP2. They were found to possess double-stranded DNA and host specificity. Morphological observations were done by electron microscopy. The bacteriophage particles stained by SYBR Green I was shown to be applicable to detect their host bacterial cells mixed with activated sludge. Two M. phosphovorus-specific bacteriophages were isolated and classified as Siphoviridae. FLPs of them were prepared, and successfully applied to detect the host bacterium added into the activated sludge. At least some of bacteria in activated sludge are susceptible to their related bacteriophages. Bacteriophages lytic to activated sludge bacteria could be affecting the bacterial population in activated sludge. The FLPs could be used for the easy-rapid detection of their host bacterium in activated sludge.

Accumulation of polyhydroxyalkanoates by Microlunatus phosphovorus under various growth conditions

Microlunatus phosphovorus is an activated-sludge bacterium with high levels of phosphorus-accumulating activity and phosphate uptake and release activities. Thus, it is an interesting model organism to study biological phosphorus removal. However, there are no studies demonstrating the polyhydroxyalkanoate (PHA) storage capability of M. phosphovorus, which is surprising for a polyphosphate-accumulating organism. This study investigates in detail the PHA storage behavior of M. phosphovorus under different growth conditions and using different carbon sources. Pure culture studies in batch-growth systems were conducted in shake-flasks and in a bioreactor, using chemically defined growth media with glucose as the sole carbon source. A batch-growth system with anaerobic-aerobic cycles and varying concentrations of glucose or acetate as the sole carbon source, similar to enhanced biological phosphorus removal processes, was also employed. The results of this study demonstrate for the first time that M. phosphovorus produces significant amounts of PHAs under various growth conditions and with different carbon sources. When the PHA productions of all cultivations were compared, poly(3-hydroxybutyrate) (PHB), the major PHA polymer, was produced at about 20-30% of the cellular dry weight. The highest PHB production was observed as 1,421 mg/l in batch-growth systems with anaerobic-aerobic cycles and at 4 g/l initial glucose concentration. In light of these key results regarding the growth physiology and PHA-production capability of M. phosphovorus, it can be concluded that this organism could be a good candidate for microbial PHA production because of its advantages of easy growth, high biomass and PHB yield on substrate and no significant production of fermentative byproducts.

Propionicimonas paludicola gen. nov., sp. nov., a novel facultatively anaerobic, Gram-positive, propionate-producing bacterium isolated from plant residue in irrigated rice-field soil

Two propionate-producing strains (Wd(T) and Wf) that were isolated anaerobically from plant residue of irrigated rice-field soil in Japan were characterized phenotypically and phylogenetically. The growth rate of strain Wd(T) was very slow in basal medium, but both growth and propionate production were stimulated significantly by the addition of cyanocobalamin. Strain Wf grew well in basal medium and produced substantial amounts of fermentation products, including propionate. Other phenotypic and phylogenetic characteristics of the two isolates were almost identical. Both were facultatively anaerobic, but much better growth was observed under anaerobic conditions. Cells were Gram-positive, non-motile, non-spore-forming and pleomorphic rods with irregular V- or crescent-shaped cell arrangements. Fermentation products from glucose in the presence of excess cyanocobalamin were acetate, lactate, a small amount of succinate and CO(2), in addition to propionate. Both oxidase and catalase activities were negative. The strains possessed meso-diaminopimelic acid in their peptidoglycan and their major cellular fatty acids were C(13 : 0), anteiso-C(15 : 0) and C(15 : 0). The isolates had high genomic DNA G+C contents (68.7 and 67.4 mol%, respectively). Menaquinones MK-9(H(4)) and MK-10(H(4)) were the predominant respiratory quinones. Phylogenetic analysis based on 16S rDNA sequences placed both strains in the Actinobacteria, with Micropruina glycogenica as their closest relative (sequence similarity values of 95.8 and 95.7 %, respectively). Microlunatus phosphovorus and Friedmanniella antarctica were also related closely to the isolates. As their morphological, physiological and chemotaxonomic characteristics were distinctly different from those of any related species, Propionicimonas paludicola gen. nov., sp. nov. is proposed to accommodate these strains. The type strain of the novel species is Wd(T) (=JCM 11933(T)=DSM 15597(T)).

Strictly polyphosphate-dependent glucokinase in a polyphosphate-accumulating bacterium, Microlunatus phosphovorus.

ATP-dependent glucokinase is suggested to have evolved from a hypothetical polyphosphate (polyP)-dependent glucokinase (polyP-GK) via a bifunctional polyP/ATP glucokinase (polyP/ATP-GK). Here we showed that polyP-GK is present in a polyP-accumulating bacterium, Microlunatus phosphovorus. The polyP-GK produced glucose-6-P(i) from glucose and polyP, but it could not phosphorylate glucose with ATP. The polyP-GK was most closely related to the polyP/ATP-GK of Mycobacterium tuberculosis.

Isolation and characterization of a Gram-positive polyphosphate-accumulating bacterium.

A Gram-positive polyphosphate-accumulating bacterium was isolated from phosphate-removal activated sludge using pyruvate-supplemented agar plates. The isolate was oval or coccobacilli (0.4-0.7 x 0.5-1.0 mm) that occurred singly, in pairs or irregular clumps. Polyphosphate granules in the cells were observed by toluidine blue staining. The pure culture of the isolate rapidly took up phosphate (9.2 mg-P/g-dry weight) in the 3-h aerobic incubation without organic substrates, after anaerobic incubation with organic substrates containing casamino acids. When acetate was the sole carbon source in the anaerobic incubation, the isolate did not remove phosphate. These physiological features of the isolate were similar to those of Microlunatus phosphovorus. However, unlike M. phosphovorus the P-removal ability of the isolate was relatively low and was not accelerated by repeating the anaerobic/aerobic incubation cycles. Phylogenetic analysis and comparison of several characteristics showed that the isolate was identified as Tetrasphaera elongata which was recently proposed as a new polyphosphate-accumulating species isolated from activated sludge. As the isolate contained menaquinone (MK)-8(H(4)) as the predominant isoprenoid ubiquinone, it may be significantly responsible for phosphate removal, because MK-8(H(4)) has reportedly been found in fairly high proportions in many phosphate-removing activated sludges.

Two exopolyphosphatases of Microlunatus phosphovorus, a polyphosphate-accumulating eubacterium from activated sludge

Two exopolyphosphatases (polyphosphatase I and II; EC 3.6.1.11) from Microlunatus phosphovorus isolated from an activated sludge, release orthophosphate from inorganic polyphosphates and have been detected and characterized for the first time. Polyphosphatase I has a molecular mass of 93 kDa, pH optimum of 4.5, does not require K + for its activity and is stimulated by divalent cations. Polyphosphatase II has a molecular mass of 55 kDa, pH optimum of 7.5 and displays its optimal activity in the presence of K + and divalent cations. The content of polyphosphatase I increases during growth, while polyphosphatase II varies only slightly.

Micropruina glycogenica gen. nov., sp. nov., a new Gram-positive glycogen-accumulating bacterium isolated from activated sludge.

A new Gram-positive non-spore-forming bacterium, strain Lg2T, was isolated from an activated sludge reactor showing enhanced biological phosphorus removal activity. The new isolate was a slowly growing organism and was capable of accumulating large amounts of intracellular glycogen from substrate taken up. Both oxidase and catalase were produced. The new isolate contained meso-diaminopimelic acid (DAP) in the cell wall. Complex fatty acid patterns with iso-C14:0, anteiso-C15:0, C16:0, iso-C16:0 and four other minor saturated or unsaturated straight-chain fatty acids were detected. The isolate contained a high genomic G+C content (70.5 mol%). Phylogenetic analysis based on the 16S rRNA gene sequence placed the isolate in the high G+C Gram-positive group with Microlunatus phosphovorus and Friedmanniella antarctica as the closest relatives (sequence similarities are 93 and 92 %, respectively). These three organisms shared common features in morphology, but strain Lg2T could be differentiated from the other species by its peptidoglycan type (meso-DAP), fatty acid composition, carbon source utilization profile and G+C content. On the basis of these findings, it is proposed that a new genus and species, Micropruina glycogenica, should be created for the new isolate; the type strain is strain Lg2T (= JCM 10248T).