Genus Leptothrix Research Articles

Study of biogenic iron oxyhydroxide for application in electronics and biotechnology

The object of this study is a biogenic material obtained from neutrophilic sheath-forming iron bacteria of the genus Leptothrix cultivated in a specific medium. Light microscopy and scanning electron microscopy were used to characterize the biogenic product. The X-ray diffraction spectrum of the control sample (without inoculated bacteria) is presented to demonstrate the important role of the bacteria in the Fe2+ transformation. Unlike the biogenic product, where nanoparticles of lepidocrocite are present, no traces of stable single-phase iron oxide or oxyhydroxide were detected in the control sample. The study of the biogenic sample showed a lack of significant differences between the naturally obtained and the artificially produced sheaths. Magnetostriction effect measurements were conducted to probe how the different magnetic behavior at temperatures below and above the Neel temperature (50 - 77 K) could be used in possible applications of the material in actuating and sensing devices.

Nepodceňujme mikrobiální precipitaci oxihydroxidů železa a manganu v životním prostředí

Dolníček (2020) described several occurrences of bog iron (limonite), which according to his article originates in recent streams by chemical precipitation, without the important role of microorganisms on this process. However, similar occurrences of reddish or orange slime, frequently accompanied by oil-like films floating on the water, are according to many papers ascribed to the biogenic iron oxidation by bacteria. To solve the origin of such precipitates, we have sampled similar materials in four places near Ostrava - Hrabová, two close to Řepiště and Staříč, and also two sites in the vicinity of Hulín investigated by Dolníček (2020). In all cases, scanning electron microscopy of dried samples at the low vacuum (ca 50 Pa) revealed a dominant share of tubular sheaths of the Leptothrix-Sphaerotilus group of bacteria. The prevailing diameter of 1.5 to 1 μm should point to genus Leptothrix, while those with a diameter up to 2.5 μm (genus Sphaerotilus) are far less common. Spiral chains of Gallionella were recognized in all samples, as well as rare pennate diatoms. Space between bacterial filaments is predominantly filled with densely stacked particles micrometer to submicrometer in size, sometimes with oval, fibrous, or tabular shape. It might be both decomposed material of previously mentioned genera and some other small-sized bacteria. Energy-dispersive spectroscopy shows dominant content of iron and oxygen, with a minor admixture of Si, Ca, Al, and P. Powder X-ray diffraction of such microbial mats is showing clastic admixtures (quartz, feldspar group minerals, illite-muscovite, and kaolinite) and two broad low-intensity peaks of ferrihydrite. Only sample D from Řepiště locality shows an absence of ferrihydrite and the presence of hematite and magnetite and/or maghemite. Therefore, we demonstrated that bacterially-induced precipitation plays a major role in the genesis of such recent bog iron occurrences and there are ways how to characterize such material by both biological and mineralogical procedures.

Study on the Composition of Biogenic Iron-Containing Materials Obtained Under Cultivation of the Leptothrix sp. on Different Media.

The biogenic iron oxide/hydroxide materials possess useful combination of physicochemical properties and are considered for application in various areas. Their production does not require special investments because these compounds are formed during cultivation of neurophilic iron bacteria. Bacteria from genus Leptothrix develop intensively in the Sphaerotilus-Leptothrix group of bacteria isolation medium and feeding media of Fedorov and Lieske. These media are different in their composition which determined the present study as an attempt to clear up the reasons that define the differences in the composition of the laboratory-obtained biomasses and the natural biomass finds. FTIRS, Mössbauer spectroscopy, and XRD were used in the research. Comparative analysis showed that the biomass and control samples contain iron compounds (α-FeOOH, γ-FeOOH, β-FeOOH, γ-Fe2O3) in different ratios. The biomass samples were enriched in oxyhydroxides of higher dispersion. Organic residuals of bacterial origin, SO4, CO3, and PO4 groups were registered in the biogenic materials.

Biogenic nanosized iron oxides obtained from cultivation of iron bacteria from the genus Leptothrix

A detailed investigation of nanostructured iron oxides/(oxy)hydroxides gathered after cultivation of bacteria from the genus Leptothrix as iron (II) oxidizers is presented. A specific type of medium is selected for the cultivation of the bacteria. Results for sediment powder and bio-film on glass substrate samples from the same media are discussed. XRD, Raman spectroscopy, SEM, and TEM images and PPMS measurements are used to prove the exact composition of the biogenic products and to interpret the oxidation process. Analysis of the data collected shows that around 80% of the iron (II) from the growth medium has been transformed into iron (III) in the form of different (oxy)hydroxides, with the rest found to be in a mixed 2,5 valence in magnetite. Our investigation shows that the bio-film sample has a phase content different from that of the powdered biomass and that lepidocrocite (γ-FeOOH) is the predominant and the initial biogenic phase in both samples. Magnetite nanoparticles are a secondary product in the bio-film, part of which possesses a defective quasi-maghemite surface layer. In the powdered biomass, the oxidation steps are not fully completed. The initial products are non-stoichiometric and due to the mixed ferric and ferrous ions present, they develop into: (i) lepidocrocite (γ-FeOOH) as a basic sediment, (ii) magnetite (Fe3O4) and (iii) goethite (α-FeOOH) in small quantities. The average size of all iron-bearing particles is found to be below 30nm. The magnetic measurements performed show a superparamagnetic behavior of the material at room temperature.

Abiotic Deposition of Fe Complexes onto Leptothrix Sheaths.

Bacteria classified in species of the genus Leptothrix produce extracellular, microtubular, Fe-encrusted sheaths. The encrustation has been previously linked to bacterial Fe oxidases, which oxidize Fe(II) to Fe(III) and/or active groups of bacterial exopolymers within sheaths to attract and bind aqueous-phase inorganics. When L. cholodnii SP-6 cells were cultured in media amended with high Fe(II) concentrations, Fe(III) precipitates visibly formed immediately after addition of Fe(II) to the medium, suggesting prompt abiotic oxidation of Fe(II) to Fe(III). Intriguingly, these precipitates were deposited onto the sheath surface of bacterial cells as the population was actively growing. When Fe(III) was added to the medium, similar precipitates formed in the medium first and were abiotically deposited onto the sheath surfaces. The precipitates in the Fe(II) medium were composed of assemblies of globular, amorphous particles (ca. 50 nm diameter), while those in the Fe(III) medium were composed of large, aggregated particles (≥3 µm diameter) with a similar amorphous structure. These precipitates also adhered to cell-free sheaths. We thus concluded that direct abiotic deposition of Fe complexes onto the sheath surface occurs independently of cellular activity in liquid media containing Fe salts, although it remains unclear how this deposition is associated with the previously proposed mechanisms (oxidation enzyme- and/or active group of organic components-involved) of Fe encrustation of the Leptothrix sheaths.

Treatment of leptothrix cells with ultrapure water poses a threat to their viability.

The genus Leptothrix, a type of Fe/Mn-oxidizing bacteria, is characterized by its formation of an extracellular and microtubular sheath. Although almost all sheaths harvested from natural aquatic environments are hollow, a few chained bacterial cells are occasionally seen within some sheaths of young stage. We previously reported that sheaths of Leptothrix sp. strain OUMS1 cultured in artificial media became hollow with aging due to spontaneous autolysis within the sheaths. In this study, we investigated environmental conditions that lead the OUMS1 cells to die. Treatment of the cells with ultrapure water or acidic buffers (pH 6.0) caused autolysis of the cells. Under these conditions, the plasma membrane and cytoplasm of cells were drastically damaged, resulting in leakage of intracellular electrolytes and relaxation of genomic DNA. The autolysis was suppressed by the presence of Ca2+. The hydrolysis of peptidoglycan by the lysozyme treatment similarly caused autolysis of the cells and was suppressed also by the presence of Ca2+. However, it remains unclear whether the acidic pH-dependent autolysis is attributable to damage of peptidoglycan. It was observed that L. discophora strain SP-6 cells also underwent autolysis when suspended in ultrapure water; it is however, uncertain whether this phenomenon is common among other members of the genus Leptothrix.

Perspectives on the Biogenesis of Iron Oxide Complexes Produced by Leptothrix, an Iron-oxidizing Bacterium and Promising Industrial Applications for their Functions

Leptothrix species, one of the Fe-/Mn-oxidizing bacteria, are ubiquitous in aqueous environments, especially at sites characterized by a circumneutral pH, an oxygen gradient and a source of reduced Fe and Mn minerals. Characteristic traits that distinguish the genus Leptothrix from other phylogenetically related species are its filamentous growth and ability to form uniquely shaped microtubular sheaths through the precipitation of copious amounts of oxidized Fe or Mn. The sheath is an ingenious hybrid of organic and inorganic materials produced through the interaction of bacterial exopolymers with aqueous-phase inorganics. Intriguingly, we discovered that Leptothrix sheaths have a variety of unexpected functions that are suitable for industrial applications such as material for lithium battery electrode, a catalyst enhancer, pottery pigment among others. This review focuses on the structural and chemical properties of the Leptothrix sheaths and their noteworthy functions that show promise for development of cost-effective, eco-friendly industrial applications.

Manganese Oxide Biominerals from Freshwater Environments in Quadrilatero Ferrifero, Minas Gerais, Brazil

Biogeochemical Mn cycling in aquatic environments is driven mostly by microbes, and includes reductive dissolution in anaerobic sediments as well as oxidation in aerobic regions. Oxidation is followed by precipitation, which occurs mainly on the extracellular structures of microorganisms. In this work, we studied the minerals precipitated on extracellular structures of native microorganisms from freshwater environments in Quadrilatero Ferrifero (Iron Quadrangle), Minas Gerais, Brazil, known to contain high levels of manganese. Light microscopy of biofilms and floating material showed diverse biomineralized structures. The most conspicuous were identified as the holdfasts of algae from the genus Ulothrix. Diatom frustules associated to manganese oxide precipitates were relatively common. In addition, both filaments and holdfasts produced by bacteria of the genus Leptothrix were found, as well as structures similar to those described as Siderocapsa and Metallogenium. Some previously unknown structures were also observed. Transmission electron microscopy of most of these structures showed the ‘crumpled tissue-paper’ morphology common in biomineralised manganese oxides. Energy-dispersive X-ray analysis (EDXA) showed that manganese and oxygen were the main components, along with minor amounts of Al, P, S, K, Ca and/or Ba. Our results bring new perspectives to the study of biomineralized manganese oxide structures from the environment. Moreover, they add information about the background of present-day microbial structures needed to better interpret fossilized microbial biominerals.

Unraveling the stratification of an iron-oxidizing microbial mat by metatranscriptomics.

A metatranscriptomic approach was used to study community gene expression in a naturally occurring iron-rich microbial mat. Total microbial community RNA was reversely transcribed and sequenced by pyrosequencing. Characterization of expressed gene sequences provided accurate and detailed information of the composition of the transcriptionally active community and revealed phylogenetic and functional stratifications within the mat. Comparison of 16S rRNA reads and delineation of OTUs showed significantly lower values of metatranscriptomic-based richness and diversity in the upper parts of the mat than in the deeper regions. Taxonomic affiliation of rRNA sequences and mRNA genome recruitments indicated that iron-oxidizing bacteria affiliated to the genus Leptothrix, dominated the community in the upper layers of the mat. Surprisingly, type I methanotrophs contributed to the majority of the sequences in the deep layers of the mat. Analysis of mRNA expression patterns showed that genes encoding the three subunits of the particulate methane monooxygenase (pmoCAB) were the most highly expressed in our dataset. These results provide strong hints that iron-oxidation and methane-oxidation occur simultaneously in microbial mats and that both groups of microorganisms are major players in the functioning of this ecosystem.

Bacterial Diversity in Biological Filtration Plant for the Removal of Iron and Manganese from Groundwater

Bacterial diversity of the microbial consortia in a biological filtration plant for the elimination of iron (Fe) and manganese (Mn) from groundwater in Joyo City, Kyoto, Japan, was studied. PCR-based denaturing gradient gel electrophoresis (DGGE) analysis of bacterial 16S ribosomal RNA (rRNA) genes represented at least 15 signals, and nucleotide sequences of the dominant fragments showed similarities to Gallionella and Nitrospira. Phylogenetic analysis using the obtained nucleotide sequences of the 16S rRNA gene clone library showed the presence of the bacteria related to Hyphomicrobium, Gallionella, and Sideroxydans, which are supposed to be involved in Fe and Mn removal. In contrast, no 16S rRNA gene clone affiliated with the genus Leptothrix, which have been regarded as a major Fe- and Mn-oxidizer in biological filtration system, was observed. Though a large number of 16S rRNA gene clones closely related to Nitrospira was obtained, no clone showed high sequence similarity to known ammonium oxidizing bacteria (AOB). However, PCR-amplification of the ammonia monooxigenase gene (amoA) of AOB and ammonia oxidizing archaea (AOA) indicated the presence of both AOB and AOA in this biological filtration plant.

Initial Assemblage of Bacterial Saccharic Fibrils and Element Deposition to Form an Immature Sheath in Cultured Leptothrix sp. Strain OUMS1

In an aquatic environment, the genus Leptothrix produces an extracellular Fe- or Mn-encrusted tubular sheath composed of a complex hybrid of bacterial exopolymers and aqueous-phase inorganic elements. This ultrastructural study investigated initial assemblage of bacterial saccharic fibrils and subsequent deposition of aqueous-phase inorganic elements to form the immature sheath skeleton of cultured Leptothrix sp. strain OUMS1. After one day of culture, a globular and/or thread-like secretion was observed on the surface of the bacterial cell envelope, and secreted bodies were transported across the intervening space away from the cell to form an immature sheath skeleton comprising assembled and intermingled fibrils. Energy dispersive X-ray microanalysis and specific Bi-staining detected a distinguishable level of P, trace Si, and a notable amount of carbohydrates in the skeleton, but not Fe. By the second day, the skeleton was prominently thickened with an inner layer of almost parallel aligned fibrils, along with low level of Fe deposition, whereas an outer intermingled fibrous layer exhibited heavy deposition of Fe along with significant deposition of P and Si. These results indicate that basic sheath-construction proceeds in two steps under culture conditions: an initial assemblage of bacterial saccharic fibrils originated from the cell envelope and the subsequent deposition of aqueous-phase Fe, P, and Si.

Bacteria associated with iron seeps in a sulfur-rich, neutral pH, freshwater ecosystem

The freshwater nature reserve De Bruuk is an iron- and sulfur-rich minerotrophic peatland containing many iron seeps and forms a suitable habitat for iron and sulfur cycle bacteria. Analysis of 16S rRNA gene-based clone libraries showed a striking correlation of the bacterial population of samples from this freshwater ecosystem with the processes of iron reduction (genus Geobacter), iron oxidation (genera Leptothrix and Gallionella) and sulfur oxidation (genus Sulfuricurvum). Results from fluorescence in situ hybridization analyses with a probe specific for the beta-1 subgroup of Proteobacteria, to which the genera Leptothrix and Gallionella belong, and newly developed probes specific for the genera Geobacter and Sulfuricurvum, supported the clone library data. Molecular data suggested members of the epsilonproteobacterial genus Sulfuricurvum as contributors to the oxidation of reduced sulfur compounds in the iron seeps of De Bruuk. In an evaluation of anaerobic dimethyl sulfide (DMS)-degrading activity of sediment, incubations with the electron acceptors sulfate, ferric iron and nitrate were performed. The fastest conversion of DMS was observed with nitrate. Further, a DMS-oxidizing, nitrate-reducing enrichment culture was established with sediment material from De Bruuk. This culture was dominated by dimorphic, prosthecate bacteria, and the 16S rRNA gene sequence obtained from this enrichment was closely affiliated with Hyphomicrobium facile, which indicates that the Hyphomicrobium species are capable of both aerobic and nitrate-driven DMS degradation.

Manganese removal and occurrence of manganese oxidizing bacteria in full-scale biofilters

Biological treatment of drinking water to remove manganese (Mn) is becoming more and more common. However, details as to how the technology works remain poorly understood. Some previous studies have focused on bacteria of the genus Leptothrix , identifying them as one of the predominant organisms in Mn oxidizing biofilms. This research took media from inside the filters of four Mn biofiltration plants in the province of New Brunswick, Canada. The water was characterized for all four sites and it was found that Mn removal from all plants is virtually 100%. Biofilm was detached from the sand and tested for presence of Leptothrix using real-tme polymerase chain reaction (RT-PCR) DNA amplification. Less specific testing for Mn oxidizing bacteria was done simultaneously using indicative agar plates. Results showed that only one plant contained Leptothrix in its filters while three of the four plants tested positive for manganese oxidizing bacteria (MOB) through plating. These results, along with other evidence, suggest that while the mechanism of oxidation observed is likely biological, Leptothrix is not necessarily the predominant organism carrying out biooxidation.

A new bacterial strain mediating As oxidation in the Fe-rich biofilm naturally growing in a groundwater Fe treatment pilot unit

A bacterial strain B2 that oxidizes arsenite into arsenate was isolated from the biofilm growing in a biological groundwater treatment process used for Fe removal. This strain is phylogenetically and morphologically different from the genus Leptothrix commonly encountered in biological iron oxidation processes. T-RFLP fingerprint of the biofilm revealed that this isolated strain B2 corresponds to the major population of the bacterial community in the biofilm. Therefore, it is probably one of the major contributors to arsenic removal in the treatment process.

LEPTOTRICHITES RESINATUS NEW GENUS AND SPECIES: A FOSSIL SHEATHED BACTERIUM IN ALPINE CRETACEOUS AMBER

Abstract Fungal hyphae, unicellular algae, and filiform prokaryotic inclusions are the most abundant microfossils of the Cretaceous amber of Schliersee (Bavaria, southern Germany). These prokaryotes are described as Leptotrichites resinatus new genus and species, and interpreted as sheathed bacteria with similarities to the extant genus Leptothrix Kutzing, 1843. However, the micromorphological and microanalytical features of this new species do not correspond entirely with those of the modern sheathed bacteria. Previous interpretations of these inclusions as filiform cyanobacteria, algae, and fungi have to be revised. Together with their numerous syninclusions, mainly fossil ciliates, testaceans, and microalgae, these prokaryotes belonged to a Cenomanian limnetic microcenosis of water bodies, such as ponds close to the resin-producing trees. Actualistic paleontological experiments reveal how these soft-bodied microorganisms could have been embedded in resins.

Some Chemotaxonomic and Physiological Properties of the Genus Sphaerotilus

Ten reference strains of the genus Sphaerotilus and five isolates of filamentous bacteria obtained from activated sludge and morphologically assigned to the genus Sphaerotilus were studied for cellular fatty acid composition, quinone type, physiological characteristics and protein profiles. All strains had the Q-8 ubiquinone as the major quinone. The main components of cellular fatty acids in all strains were the straight chain unsaturated acid 16:1 and saturated acid 16:0. All strains contained the hydroxylated fatty acid 3-OH 10:0, sometimes only in minor amounts or traces. Analysis of physiological properties showed phenotypic homogeneity among the tested organisms. From 240 tested organic compounds, several sugars, organic acids and aromatic compounds could be utilized as sole carbon source. A clear differentiation from strains belonging to the morphological similar genus Leptothrix was possible with several carbon source utilization tests. Standardized protein profiles of the isolates were also very similar indicating a high degree of homogeneity. The results of the chemotaxonomic characterization are in line with the phylogenetic allocation of the genus Sphaerotilus within the Rubrivivax subdivison of the β subclass of the Proteobacteria .

Low-temperature hydrothermal deposits of Franklin Seamount, Woodlark Basin, Papua New Guinea

Hydrothermal deposits of siliceous FeMn oxides from a few cm to 7 m in height are widespread at the summit and caldera floor of Franklin Seamount, a 250 m high edifice of basaltic andesite lava tubes, pillows and talus located near the western propagating tip of the Woodlark spreading axis. Some deposits are venting clear fluid at about 30 °C. The hydrothermal constructions are composed predominantly of X-ray amorphous protoferrihydrite mixed with variable amounts of opaline silica and greenish nontronite. Most deposits are covered by mm-thick black crusts of vernadite and feroxyhyte which are mineralised bacterial formations (genus Leptothrix) of hydrogenous origin. Microbes also influenced formation of the Fe-rich parts of the deposits, as evidenced by filamentous protoferrihydrite pseudomorphs of Gallionella. Blocky inorganic particles of protoferrihydrite and ferrihydrite occur as rare interior phases. Nontronite formed by precipitation from hydrothermal solutions, commonly involving replacement of microbial protoferrihydrite and silica. Patchy and lenticular birnessite, partly altered by biogenic processes to vernadite, occurs within siliceous protoferrihydrite matrices in dark cm-thick outer margins of some edifices. This is considered hydrothermal in origin, reflecting percolation of hydrothermal fluids through porous protoferrihydrite-opal aggregates to more oxidised conditions near the exterior related to mixing with seawater. The different mineralogical associations are reflected by geochemistry. Trace elements in the manganiferous phases were derived partly from the primary hydrothermal solution and partly from seawater with which this was extensively diluted at the depositional site and also deeper within the volcanic structure of Franklin Seamount. Most of the Fe, Mn, Cu, Zn and Pb was derived from the primary hydrothermal fluid. From limited data, nontronitic portions of the deposits contain more Cu, Zn and Pb than the siliceous protoferrihydrite-rich portions. Migration of redox boundaries and redistribution of elements during flow-through processes influence development of the typical zoned structure of the deposits. Uranium-thorium decay series equilibria at both actively venting and inactive sites at Franklin Seamount yield ages not exceeding 43 years.

Polyphasic Characterization of the Genus Leptothrix: New Descriptions of Leptothrix mobilis sp. nov. and Leptothrix discophora sp. nov. nom. rev. and Emended Description of Leptothrix cholodnii emend.

Strain Feox-1 was isolated from the sediment of a big freshwater lake (Chiemsee) in Upper Bavaria, Germany. The site of isolation was characterized by dark-brown ferromanganese deposits produced by sheated bacteria belonging to the genus Leptothirix. Cells of the newly isolated strain were gram-negative, rodshaped and highly motile by a single polar flagellum. Although sheaths were not formed strain Feox-1 could be affiliated to the genus Leptothrix by its ability to oxidize manganese at circumneutral pH values. A polyphasic approach was used to determine the taxonomic status of Feox-1 and of the other currently available Leptothrix strains which have been only partly described. Physiological tests, fatty acid analyses, DNA-DNA hybridization experiments and sequencing of 16S rRNA genes indicated that Feox-1 belongs to a new species, for which the name Leptothrix mobilis is proposed. Main characteristics of the newly proposed species are as follows: Good growth occurs in the range of 10 to 37°C and pH 6.5 to 8.5. Only very few compounds are utilized as sole carbon source. Cells contain as major fatty acids hexadecanoic acid, cis-9-hexadecenoic acid and octadecenoic acid; minor fatty acids are 3-hydroxydecanoic acid, dodecanoic acid, tetradecanoic acid and cis-9,10-methyleneoctadecanoic acid. The guanine-plus-cytosine content of the DNA is 68 mol%. Taxonomic studies of strains belonging to the non-validly described species “Leptothrix discophora” and to Leptottirix cbolodnii led to the description of Leptottirix discophora sp. nov. nom. rev. and to an emended description of Leptottirix cbolodnii. In addition, the two strains LMG 8142 and LMG 8143 originally assigned to "Leptottirix discophora" were transferred to Leptothrix cbolodnii.

Phylogeny of the Sphaerotilus-Leptothrix group inferred from morphological comparisons, genomic fingerprinting, and 16S ribosomal DNA sequence analyses.

Phase-contrast light microscopy revealed that only one of eight cultivated strains belonging to the Sphaerotilus-Leptothrix group of sheathed bacteria actually produced a sheath in standard growth media. Two Sphaerotilus natans strains produced branched cells, but other morphological characteristics that were used to identify these bacteria were consistent with previously published descriptions. Genomic fingerprints, which were obtained by performing PCR amplification with primers corresponding to enterobacterial repetitive intergenic consensus sequences, were useful for distinguishing between the genera Sphaerotilus and Leptothrix, as well as among individual strains. The complete 16S ribosomal DNA (rDNA) sequences of two strains of "Leptothrix discophora" (strains SP-6 and SS-1) were determined. In addition, partial sequences (approximately 300 nucleotides) of one strain of Leptothrix cholodnii (strain LMG 7171), an unidentified Leptothrix strain (strain NC-1), and four strains of Sphaerotilus natans (strains ATCC 13338T [T = type strain], ATCC 15291, ATCC 29329, and ATCC 29330) were determined. We found that two of the S. natans strains (ATCC 15291 and ATCC 13338T), which differed in morphology and in their genomic fingerprints, had identical sequences in the 300-nucleotide region sequenced. Both parsimony and distance matrix methods were used to infer the evolutionary relationships of the eight strains in a comparison of the 16S rDNA sequences of these organisms with 16S rDNA sequences obtained from ribosomal sequence databases. All of the strains clustered in the Rubrivivax subdivision of the beta subclass of the Proteobacteria, which confirmed previously published conclusions concerning selected individual strains. Additional analyses revealed that all of the S. natans strains clustered in one closely related group, while the Leptothrix strains clustered in two separate lineages that were approximately equidistant from the S. natans cluster. This finding suggests that the tentative species "L. discophora" needs to be more clearly defined and compared with other species belonging to the genus Leptothrix.

On the “ Muffa “ of the Sulphur Springs at Valdieri

ABSTRACT The baths of Valdieri, not far from a Piedmontese town of that name, are situated in a valley on the northern side of the Maritime Alps, and have long been celebrated, not only for the coolness of their climate and the excellence of their mineral waters, but also for the “Muffa,” a sub- stance occurring in one of those waters, which, while of great medicinal value as an external application, is interesting, when viewed under the microscope, for the vegetable, animal, and mineral productions which it contains. These baths are 4426 feet above the level of the sea. Found in those sulphur springs which have a temperature of about fifty degrees Centigrade, the Muffa first appears as tender minute filaments, soft and floating, of a greenish-white colour, surrounded by a mucilaginous milky-white substance imbued with a sulphurous deposit. Of little consistency in its early state, it soon becomes more substantial; changing in colour to violet, then light yellow, and finally to a pale green. When mature, the Muffa resembles a gelatinous lard, carpeting the rock down which the water flows. The vegetable above referred to was considered by Allioni to be Ulva labyrinthiformis of Linnæus. In 1837 Fontan detected a distinct organization, describing it as composed of white filaments from to of a millimetre in diameter; tubular, cylindrical, simple, devoid of septa, containing small semi-opaque globules, collocated when young, and separated towards the ends of the tubes in mature individuals. To this plant he gave the name Sulphuraria, it not having been found in any except sulphur springs. Delponte, of the Botanic Garden at Turin, after careful microscopic examination, places it in the genus Leptothrix (Kützing), near to L. compacta and L. lamellosa, naming it after the place of its nativity, Valderia. A parasitic Ulva accompanies the above, growing upon it, and an Oscillatoria sometimes covers the upper surface, where the water has not more than thirty degrees of temperature. Conferva nigra also occurs.