Chlorobium chlorochromatii sp. nov., a symbiotic green sulfur bacterium isolated from the phototrophic consortium “Chlorochromatium aggregatum”

A symbiotic green sulfur bacterium, strain CaD, was isolated from an enrichment culture of the phototrophic consortium “Chlorochromatium aggregatum”. The capability of the epibiont to grow in pure culture indicates that it is not obligately symbiotic. Cells are Gram-negative, nonmotile, rod-shaped and contain chlorosomes. Strain CaD is obligately anaerobic and photolithoautotrophic, using sulfide as electron donor. Acetate and peptone are photoassimilated in the presence of sulfide and hydrogencarbonate. Photosynthetic pigments contain bacteriochlorophylls a and c, and γ-carotene and OH-γ-carotene glucoside laurate as the dominant carotenoids. In cells from pure cultures, chlorosomes are equally distributed along the inner face of the cytoplasmic membrane. In contrast, the distribution of the chlorosomes in symbiotic epibiont cells is uneven, with chlorosomes being entirely absent at the site of attachment to the central bacterium. The symbiotic epibiont cells display a conspicuous additional layered structure at the attachment site. The G + C content of genomic DNA of strain CaD is 46.7 mol%. On the basis of 16S rRNA sequence comparison, the strain is distantly related to Chlorobium species within the green sulfur bacteria phylum (≤94.6% sequence homology). The novel isolate is therefore described as a novel species within the genus Chlorobium, Chlorobium chlorochromatii.     

<Emphasis Type="Italic">Chlorobaculum macestae</Emphasis> sp. nov., a new green sulfur bacterium

The investigated green sulfur bacterium, strain M, was isolated from a sulfidic spring on the Black Sea Coast of the Caucasus. The cells of strain M are straight or curved rods 0.6–0.9 × 1.8–4.2 μm in size. According to the cell wall structure, the bacteria are gram-negative. Chlorosomes are located along the cell periphery. Strain M is an obligate anaerobe capable of photoautotrophic growth on sulfide, thiosulfate, and H₂. Acetatate is utilized as an additional carbon source. It utilizes ammonium, urea, casein hydrolysate, and N₂ as nitrogen sources and sulfide, thiosulfate, and elemental sulfur as sulfur sources. Bacteriochlorophyll c and the carotenoid chlorobactene are the main pigments. The optimal growth temperature is 25–28°C; the optimal pH is 6.8. The strain does not require NaCl. Vitamin B 12 stimulates growth. The content of the G+C base pairs in the DNA of strain M is 58.3 mol %. In the phylogenetic tree constructed on the basis of analysis of nucleotide sequences of 16S rRNA genes, strain M forms a separate branch, which occupies an intermediate position between the phylogenetic cluster containing representatives of the genus Chlorobaculum (94.9–96.8%) and the cluster containing species of the genus Chlorobium (94.1–96.5%). According to the results of analysis of the amino acid sequence corresponding to the fmo gene, strain M represents a branch which, unlike that in the “ribosomal” tree, falls into the cluster of the genus Chlorobaculum (95.8–97.2%). Phylogenetic analysis of the amino acid sequence corresponding to the nifH gene placed species of the genera Chlorobaculum and Chlorobium into a single cluster, whereas strain M formed a separate branch. The results obtained allow us to describe strain M as a new species of the genus ChlorobacChlorobaculum — Chlorobaculum macestae sp. nov.     

Growth promotion of Chlorella ellipsoidea by co-inoculation with Brevundimonas sp. isolated from the microalga

Eight bacterial strains identified as P1, P2, Y1, Y2, W1, W2, G, and R were isolated from a long-term laboratory culture of the green alga Chlorella ellipsoidea. Although it is unknown how these bacterial strains have been maintained with the C. ellipsoidea culture, all appeared to promote the growth of C. ellipsoidea. Co-inoculation of each bacterial strain with C. ellipsoidea resulted in 0.5–3 times greater algal growth than that of C. ellipsoidea alone. The most effective bacterium (i.e., strain P1) was selected and further characterized. Biochemical analysis and transmission electron microscopy revealed that strain P1 is closely related to the genus Brevundimonas. Sequence analysis of the 16S rRNA of strain P1 showed 99.9 and 99.4% nucleotide sequence identity to that of B. nasdae and B. vesicularis, respectively. In addition to the growth promotion of C. ellipsoidea by strain P1, the growth of strain P1 was also significantly enhanced by co-culturing with C. ellipsoidea, indicating a symbiotic relationship between the bacterium and alga. Scanning electron microscopy showed the direct adhesion of strain P1 cells to the surface of C. ellipsoidea cells, as well as the development of abundant crinkles on the surface of co-cultured C. ellipsoidea cells. Handling editor: J. Padisak     

Characterization of a novel plant growth-promoting bacteria strain Delftia tsuruhatensis HR4 both as a diazotroph and a potential biocontrol agent against various plant pathogens

A novel, plant growth-promoting bacterium Delftia tsuruhatensis, strain HR4, was isolated from the rhizoplane of rice (Oryza sativa L., cv. Yueguang) in North China. In vitro antagonistic assay showed this strain could suppress the growth of various plant pathogens effectively, especially the three main rice pathogens (Xanthomonas oryzae pv. oryzae, Rhizoctonia solani and Pyricularia oryzae Cavara). Treated with strain HR4 culture, rice blast, rice bacterial blight and rice sheath blight for cv. Yuefu and cv. Nonghu 6 were evidently controlled in the greenhouse. Strain HR4 also showed a high nitrogen-fixing activity in N-free Döbereiner culture medium. The acetylene reduction activity and ¹⁵N₂-fixing activity (N₂FA) were 13.06 C₂H₄ nmol ml⁻¹ h⁻¹ and 2.052 ¹⁵Na.e.%, respectively. The nif gene was located in the chromosome of this strain. Based on phenotypic, physiological, biochemical and phylogenetic studies, strain HR4 could be classified as a member of D. tsuruhatensis. However, comparisons of characteristics with other known species of the genus Delftia suggested that strain HR4 was a novel dizotrophic PGPB strain.     

Draft genome sequence of marine sediment-derived red pigmented bacteria Zooshikella sp. strain S2.1 with potential biomedical applications

The present study is aimed to determine the draft genome of novel species of Zooshikella strain S2.1, a potential red pigmented strain isolated recently from the coastal sediment of Andaman and Nicobar Islands, India. This Gram negative, rod shaped aerobic bacterium produces pink, yellowish-red and dark red with metallic green sheen pigmentation on agar plates. It is able to grow under NaCl concentrations of 1 to 9%. This species has antimicrobial, antioxidant, dye and food colorant applications. Whole genome sequence analysis revealed that strain S2.1 represents a novel species of the genus Zooshikella. Draft genome and 16 s rRNA sequences of this species were deposited in GenBank under the Sequence Read Archive accession number PRJNA514840 and GenBank number MK680108, respectively. Here we report the draft genome of Zooshikella sp. strain S2.1 with ~5.9 Mb of chromosomal content and ~0.34 Mb of extra-chromosomal content.     

Development of gene transfer methods forRubrivivax gelatinosus S1: construction,characterization and complementation of apuf operon deletion strain

Gene transfer systems were developed inRubrivivax (Rx.) gelatinosus S1. First, a system for conjugative transfer of mobilizable plasmids fromEscherichia coli toRx. gelatinosus S1 was established. Secondly, optimal conditions for the transformation ofRx. gelatinosus S1 by electroporation were determined. A Δpuf strain was constructed. Complementation with thepuf operon from a wild-type strain cloned in a replicative plasmid restored photosynthetic growth. Two insertion strains were also selected. All the strains constructed were green, due to a change in carotenoid content. Characterization of these strains provides genetic evidence for a “superoperon” organization in this bacterium.     

Whole-Genome Sequence of N-Acylhomoserine Lactone-Synthesizing and -Degrading Acinetobacter sp. Strain GG2

Acinetobacter sp. strain GG2 is a quorum-sensing and quorum-quenching bacterium isolated from the ginger rhizosphere. It degrades a broad range of N-acylhomoserine lactone molecules via lactonase. The genome sequence of strain GG2 may provide insights on the regulation of quorum-sensing and quorum-quenching mechanisms in this bacterium.     

A potential bacterial biocontrol agent,strain S2V2 against pathogenic marine <Emphasis Type="Italic">Vibrio</Emphasis> in aquaculture

We isolated a marine bacterium strain S2V2 which inhibited the growth of pathogenic marine Vibrio spp. The aims of this research were to identify a new antibiotic-producing marine bacterium strain S2V2, and evaluate its spectrum activity and pathogenic property. Analysis of 16S rDNA sequence placed strain S2V2 in the genus Pseudoalteromonas, but the sequence similarity was low (95.46%) implying the strain might be a new species in this genus. Strain S2V2 inhibited the growth of 67.9% of 28 Vibrio strains tested. This strain inhibited V. alginolyticus, V. anguillarum, V. fluvialis, V. harveyi, V. metschnikovii, V. splendidus, V. ordalii, V. parahaemolyticus, and V. vulnificus, but inactive against V. campbellii, Aeromonas hydrophyla and Staphylococcus aureus. Strain S2V2 produced extracellular non proteinaceous antibacterial substances. The highest antibacterial activity was found when strain S2V2 was cultured for 96 h in ZoBell broth medium. An artificial infection to post larvae of Lithopenaeus vanname indicated that strain S2V2 was a non pathogenic bacterium. Non pathogenic property and specific antibacterial activity against a broad range of fish pathogenic marine Vibrio of strain S2V2 suggest that this strain is a prospective source of unique antibiotic and a potential biocontrol agent in marine aquaculture.     

Occurrence of Hydrogenases in Cyanobacteria and Anoxygenic Photosynthetic Bacteria: Implications for the Phylogenetic Origin of Cyanobacterial and Algal Hydrogenases

Hydrogenases are important enzymes in the energy metabolism of microorganisms. Therefore, they are widespread in prokaryotes. We analyzed the occurrence of hydrogenases in cyanobacteria and deduced a FeFe-hydrogenase in three different heliobacterial strains. This allowed the first phylogenetic analysis of the hydrogenases of all five major groups of photosynthetic bacteria (heliobacteria, green nonsulfur bacteria, green sulfur bacteria, photosynthetic proteobacteria, and cyanobacteria). In the case of both hydrogenases found in cyanobacteria (uptake and bidirectional), the green nonsulfur bacterium Chloroflexus aurantiacus was found to be the closest ancestor. Apart from a close relation between the archaebacterial and the green sulfur bacterial sulfhydrogenase, we could not find any evidence for horizontal gene transfer. Therefore, it would be most parsimonious if a Chloroflexus-like bacterium was the ancestor of Chloroflexus aurantiacus and cyanobacteria. After having transmitted both hydrogenase genes vertically to the different cyanobacterial species, either no, one, or both enzymes were lost, thus producing the current distribution. Our data and the available data from the literature on the occurrence of cyanobacterial hydrogenases show that the cyanobacterial uptake hydrogenase is strictly linked to the occurrence of the nitrogenase. Nevertheless, we did identify a nitrogen-fixing Synechococcus strain without an uptake hydrogenase. Since we could not find genes of a FeFe-hydrogenase in any of the tested cyanobacteria, although strains performing anoxygenic photosynthesis were also included in the analysis, a cyanobacterial origin of the contemporary FeFe-hydrogenase of algal plastids seems unlikely. Electronic Supplementary Material Electronic Supplementary material is available for this article at and accessible for authorised users. [Reviewing Editor: Dr. Lauren Ancel Meyers]     

Irradiance Effects on Growth and Bacteriochlorophyll Content of Phototrophic Heliobacteria,Purple and Green Photosynthetic Bacteria

Two species of heliobacteria along with a purple and green bacterium were tested for their ability to grow phototrophically at irradiances ranging from 0.125 to 50 W m^–2. The heliobacteria were incapable of growth below 0.5 W m^–2, while both the purple and green bacterium grew at significantly lower irradiances. Specific bacteriochlorophyll contents were higher for the purple and green bacteria than for the heliobacteria at all irradiances tested. Thus in distinct contrast to purple and green bacteria, heliobacteria are high-irradiance phototrophs, and this characteristic may influence their distribution in nature.     

Spread and change in stress resistance of Shiga toxin-producing Escherichia coli?O157 on fungal colonies

To elucidate the effect of fungal hyphae on the behaviour of Shiga toxin-producing Escherichia coli (STEC) O157, the spread and change in stress resistance of the bacterium were evaluated after coculture with 11 species of food-related fungi including fermentation starters. Spread distances of STEC O157 varied depending on the co-cultured fungal species, and the motile bacterial strain spread for longer distances than the non-motile strain. The population of STEC O157 increased when co-cultured on colonies of nine fungal species but decreased on colonies of Emericella nidulans and Aspergillus ochraceus. Confocal scanning microscopy visualization of green fluorescent protein-tagged STEC O157 on fungal hyphae revealed that the bacterium colonized in the water film that existed on and between hyphae. To investigate the physiological changes in STEC O157 caused by co-culturing with fungi, the bacterium was harvested after 7 days of co-culturing and tested for acid resistance. After co-culture with eight fungal species, STEC O157 showed greater acid resistance compared to those cultured without fungi. Our results indicate that fungal hyphae can spread the contamination of STEC O157 and can also enhance the stress resistance of the bacteria.     

The structure of the 5 S ribosomal RNA of a member of the phylum of green non-sulfur bacteria and relatives

The 5 S rRNA sequence was determined for the bacterium Herpetosiphon strain Senghas Wie 2. It is the first 5 S RNA sequence reported for a member of the eubacterial phylum defined by green non-sulfur bacteria. The sequence fits into a consensus secondary structure model for eubacterial 5 S RNA. At four positions, the sequence shows substitutions with respect to strongly conserved nucleotides found in other hitherto examined eubacterial 5 S RNAs.     

Novel halotolerant bacterium from cryopeg in permafrost: Description of <Emphasis Type="Italic">Psychrobacter muriicola</Emphasis> sp. nov.

A novel halotolerant psychrotrophic gram-negative bacterium, strain 2pS, was isolated from lenses of water brine in Arctic permafrost (cryopeg). The optimal growth of the new strain was observed at 16–18°C; the maximal and minimal growth temperatures were 37°C and ?2°C, respectively. The pH growth range was 5.8 to 8.5 (optimum 6.5–7.5) and the range of medium salinity was 0 to 100 g/l (optimum 3–8 g/l NaCl). The strain 2pS did not produce acid from carbohydrates and utilized acetate, yeast extract, pyruvate, glutarate, fumarate, caproate, heptanoate, butyrate, malate, DL-lactate, citrate, L-proline, L-tyrosine, butanol, and dulcitol as the sole carbon and energy sources. The major fatty acids of the cell wall at optimal growth temperature were C_18:1ω7 and C_18:1ω9. The G+C DNA base content was 46.0 mol.%. Phylogenetic analysis of the 16S rRNA gene sequences showed that the studied strain was the closest (97% similarity) to Psychrobacter nivimaris DSM 16093^T, a halotolerant psychrotrophic bacterium isolated from the Arctic sea’s ice. Genotypic and phenotypic differences of the new bacterium from closely related species lead to the conclusion that strain 2pS belongs to a novel species of the genus Psychrobacter: Psychrobacter muriicola sp. nov.     

Nitrogen-fixing cyanobacterium <Emphasis Type="Italic">Trichormus variabilis</Emphasis> of the Lake Baikal phytoplankton

A new filamentous cyanobacterial strain BAC 9610 was isolated from the lake Baikal pelagial. Data obtained by light, scanning, and transmission electron microscopy, along with 16S rRNA gene sequence analysis, allowed the bacterium identification as Trichormus variabilis, previously known as Anabaena variabilis. Trichormus is a cyanobacterial genus not presented in the list of Baikal plankton algae; A. variabilis also hasn’t been previously detected in Baikal phytoplankton. T. variabilis nitrogen fixation ability was demonstrated. The gene responsible for nitrogen fixation, nifH, was identified by PCR and was partially sequenced. No hepatotoxin synthesis genes were revealed in the strain.     

Isolation and characterization of a unicellular manganese-oxidizing bacterium from a freshwater lake in northwestern Russia

A unicellular manganese-oxidizing bacterium (strain L7), isolated from Lake Ladoga, is identified as “Siderocapsa” sp. according to its morphology. However, this bacterium belongs to the phylogenetic cluster of Pseudomonas putida. The physiological characteristics (utilization of sugars, polyols, organic acids, and phenolic substrates as carbon and energy sources) also indicate the similarity of strain L7 to representatives of the genus Pseudomonas. The growing culture oxidizes Mn(II); the rate of oxidation depends on the type of added organic substrate. Carbonate requirement for this process indicates mixotrophic metabolism. The relatedness of the isolated bacterium to the representatives of the genus Pseudomonas and their phenotypic similarity provide a basis for considering strain L7 not as “Siderocapsa” sp., but as a new species, Pseudomonas siderocapsa sp. nov., of the P. putida cluster.     

Gene Deletions Resulting in Increased Nitrogen Release by Azotobacter vinelandii: Application of a Novel Nitrogen Biosensor

Azotobacter vinelandii is a widely studied model diazotrophic (nitrogen-fixing) bacterium and also an obligate aerobe, differentiating it from many other diazotrophs that require environments low in oxygen for the function of the nitrogenase. As a free-living bacterium, A. vinelandii has evolved enzymes and transporters to minimize the loss of fixed nitrogen to the surrounding environment. In this study, we pursued efforts to target specific enzymes and further developed screens to identify individual colonies of A. vinelandii producing elevated levels of extracellular nitrogen. Targeted deletions were done to convert urea into a terminal product by disrupting the urease genes that influence the ability of A. vinelandii to recycle the urea nitrogen within the cell. Construction of a nitrogen biosensor strain was done to rapidly screen several thousand colonies disrupted by transposon insertional mutagenesis to identify strains with increased extracellular nitrogen production. Several disruptions were identified in the ammonium transporter gene amtB that resulted in the production of sufficient levels of extracellular nitrogen to support the growth of the biosensor strain. Further studies substituting the biosensor strain with the green alga Chlorella sorokiniana confirmed that levels of nitrogen produced were sufficient to support the growth of this organism when the medium was supplemented with sufficient sucrose to support the growth of the A. vinelandii in coculture. The nature and quantities of nitrogen released by urease and amtB disruptions were further compared to strains reported in previous efforts that altered the nifLA regulatory system to produce elevated levels of ammonium. These results reveal alternative approaches that can be used in various combinations to yield new strains that might have further application in biofertilizer schemes.     

Degradation of Microcystin-RR by Sphingomonas sp. CBA4 isolated from San Roque reservoir (Córdoba – Argentina)

We report the aerobic biodegradation of Microcystin-RR (MC-RR) by a bacterial strain isolated from San Roque reservoir (Córdoba – Argentina). This bacterium was identified as Sphingomonas sp. (CBA4) on the basis of 16S rDNA sequencing. The isolated strain was capable of degrading completely MC-RR (200 μg l⁻¹) within 36 h. We have found evidence that MC-RR biodegradation pathway by this Sphingomonas sp. strain would start by demethylating MC-RR, affording an intermediate product, which is finally biodegraded by this strain within 72 h. Our results confirm that certain environmental bacteria, living in the same habitat as toxic cyanobacteria, have the capability to perform complete biodegradation of MC, leading to natural bioremediation of waterbodies. The bacterium reported here presents genetic homologies with other strains that degrade MC-LR. However, initial demethylation of MC-RR has been not described previously, raising questions on the probable presence of different biodegradation pathways for different MC variants.     

Photoproduction of H2 from acetate by syntrophic cocultures of green sulfur bacteria and sulfur-reducing bacteria

The marine green sulfur bacterium Chlorobium vibrioforme strain 1930 produced H₂ and elemental sulfur from sulfide or thiosulfate under N limitation in the light. H₂ production depended on nitrogenase and occurred only in the absence of ammonia. Methionine sulfoximine, an inhibitor of glutamine synthetase, prevented the switch-off by ammonia. In defined syntrophic cocultures of the acetate-oxidizing, sulfur-reducing bacterium Desulfuromonas acetoxidans with green sulfur bacteria, H₂ was produced from acetate via a light-driven sulfur cycle. The sulfur-reducing bacterium could not be replaced by sulfate-reducing bacteria in these experiments. In a coculture of the marine Chlorobium vibrioforme strain 1930 and the sulfur-reducing bacterium Desulfuromonas acetoxidans strain 5071, optimum long-term H₂ production from acetate was obtained with molecular nitrogen as N source, at low light intensity (110 mol · m^-2 · s^-1), in sulfide-reduced mineral medium (2 mM Na₂S) at pH 6.8. Traces of sulfide (10 M) were sufficient to keep the sulfur cycle running. The coculture formed no poly--hydroxyalkanoates (PHA), but 20%–40% polysaccharide per cell dry mass. Per mol acetate added, the coculture formed 3.1 mol of H₂ (78% of the theoretical maximum). Only 8% of the reducing equivalents was incorporated into biomass. The maximum rate of H₂ production was 1300 ml H₂ per day and g cell dry mass.Non-standard abbrevations MOPS 2-(N-morpholino) propane sulfonic acid - MSX Methionine sulfoximine - PHA poly--hydroxyalkanoates     

Identification of an anaerobic bacterium which reduces perchlorate and chlorate asWolinella succinogenes

A bacterium coded as strain HAP-1 was isolated from a municipal anaerobic digestor for its ability to reduce >7000 ppm perchlorate in wastewaters. The organism is capable of the dissimilatory reduction of perchlorate on chlorate to chloride for energy and growth. It is a Gram-negative, non-sporeforming, obligately anaerobic, motile thin rod. Antibiotic resistance, utilization of carbon substrates and utilization of electron acceptors by bacterium HAP-1 were similar toWolinella succinogenes. The organism's 16S rRNA sequence was 0.75% different from that of the type strain ofW. succinogenes. The fatty acid compositions of the two organisms are very similar. The morphological, physiological and 16S rRNA sequence data indicated that bacterium HAP-1 is a strain ofW. succinogenes that can utilize perchlorate or chlorate as a terminal electron acceptor.     

A phototrophic gliding filamentous bacterium of hot springs,Chloroflexus aurantiacus,gen. and sp. nov.

Chloroflexus aurantiacus, gen. and sp. n., is a filamentous phototrophic bacterium of hot springs. On an agar surface, holotype strain J-10-fl glides at 0.01–0.04 μm/sec. The filaments are 0.6–0.7 μm in width and indeterminate in length. Pigments include bacteriochlorophyll c and bacteriochlorophyll a (identified by spectrophotometry) in addition to β and γ-carotene and glycosides of the latter. Chlorobium vesicles are present. Photoheterotrophic growth occurs under anaerobic conditions. Aerobic chemoheterotrophic growth also occurs in darkness or light. Bacteriochlorophyll syntheses cease under aerobic conditions but some types of carotenoids continue to be made. The filament coloration is orange under all except anaerobic conditions in low light intensity where it is dull green. The pH optimum is near 8, the temperature optimum between 52° and 60°C. The DNA base composition for strain J-10-fl is 54.9 ± 1.0 moles % guanine + cytosine. Chloroflexus is unique in that there have been no previous reports of filamentous or gliding phototrophic bacteria. The combinations of bacteriochlorophylls a and c and the presence of chlorobium vesicles in a photoheterotroph and in an organism capable of aerobic growth are also unique. This metabolically versatile organism extends the taxonomic and phylogenetic limits of the “green line” of phototrophic bacteria.