A modified method for the cultivation of phototrophic bacteria

Simplified anaerobic media and a convenient method for the cultivation of Rhodospirillaceae, Chlorobiaceae, Chloroflexaceae and Chromatiaceae are described. The modified conditions assure almost complete anaerobiosis for media, growth and maintenance.Strains representing several species of Rhodospirillaceae, Chlorobiaceae and Chromatiaceae were successfully grown within relatively short times with full pigmentation, indicating that the new media and cultivation conditions were most suited for photoautotrophic growth.     

Occurrence and dynamics of phototrophic purple nonsulphur bacteria compared with other asymbiotic nitrogen fixers in ricefields of Egypt

The occurrence and the dynamics of phototrophic purple nonsulphur bacteria (PPNSB) as well as Azospirillum, Azotobacter, Clostridium, and cyanobacteria at different rice growth stages were studied in two ricefields, at Kafr-El-Shiekh and Al-Fayoum in Egypt.The PPNSB existed in the both rice fields examined, but their numbers varied according to field conditions, habitat and rice growth stage. After transplanting, the number of PPNSB increased gradually, reached its maximum at maximum tillering stage, and thereafter declined toward harvest time. Numbers of PPNSB were generally comparable with that of the heterotrophic N₂-fixers namely Azospirillum, Azotobacter, Clostridium and cyanobacteria, while that of phototrophic purple and green sulphur bacteria were relatively lower.The highest PPNSB numbers were generally found in rhizosphere (10³–10⁶ per g^–1 dw soil) followed by soil (10³–10⁵ per g^–1 dw soil) and floodwater (10–10² per ml). Rice plants showed a positive rhizosphere effect on PPNSB, clostridia, Azotobacter and Azospirillum, negative rhizosphere effect on cyanobacteria and green sulphur bacteria, and no effect on purple sulphur bacteria.     

ELISA analyses of malate dehydrogenases from nonsulfur purple phototrophic bacteria

Abstract The accumulation of ppGpp in three streptococci starved for isoleucine was studied via HPLC analysis of cell extracts prepared from mechanically disrupted bacteria. Starvation was achieved either by reduction of isoleucine in the growth medium or the addition of pseudomonic acid. The results indicate that while both treatments produced a physiological response similar to that described for stringent strains of other bacteria, in the streptococci, stringency was not necessarily coupled with ppGpp.     

Cryopreservation of anoxygenic phototrophic Fe(II)-oxidizing bacteria

Preservation and storage of microbial stock cultures is desirable since the risk of contamination or loss of living cultures is immanent while over long periods mutations accumulate. Generally, it is rather difficult to preserve photosynthetic bacteria due to their sensitive photosynthetic membranes [1]. Phototrophic Fe(II)-oxidizing bacteria face an additional challenge; since they are exposed to light and Fe(II) during growth, they have to cope with radicals from Fenton reactions of Fe(II)-species, light and water. Therefore, phototrophic Fe(II)-oxidizing strains are thought to be especially susceptible to genetic modifications. Here, we provide a simple and fast protocol using glycerol as cryo-protectant to cryopreserve three strains of anoxygenic phototrophic Fe(II)-oxidizing bacteria from different taxa: α-proteobacteria, γ-proteobacteria and chloroflexi. All three strains investigated could be revived after 17 months at −72 °C. This suggests that a long-term storage of phototrophic Fe(II)-oxidizing strains is possible.     

A new method for liquid nitrogen storage of phototrophic bacteria under anaerobic conditions

A simple, effective and economical method for the long-term preservation of bacteria in liquid nitrogen under anaerobic conditions is described. As a case example anaerobic photosynthetic bacteria were successfully preserved. Gas tight small screw-cap glass ampoules with butyl rubber septa were used for freezing the specimen anaerobically. During experimental manipulations no anaerobic chamber or glove boxes were required. All teste cultures yielded high recoveries after repeated thawing and during storage. After freezing, survival recoveries of Rhodospirillaceae range from 70–100%, whereas with strict anaerobic strains of Chlorobiaceae and Chromatiaceae a maximum loss of 1–2 log₁₀ counts was observed. No further loss in viability occurred after 1–2 years of storage.The small size of the ampoules and the use of single ampoule for 15–20 repeated retrievals proved economical with respect to storage space and costs.The system is compact and suitable for the preservation of anaerobic phototropic bacteria and other fragile anaerobic microorganisms.     

Cysteine and S-sulfocysteine biosynthesis in phototrophic bacteria

Forteen species (17 strains) of phototrophic bacteria as well as one strain of Thiobacillus denitrificans were tested for cysteine synthase and S-sulfocysteine synthase. All strains contain cysteine synthase active with O-acetylserine; only the Chromatiaceae, two species of the Rhodospirillaceae and T. denitrificans contain S-sulfocysteine synthase. In six species repression by different sulfur compounds in the medium was studied. In Chromatium vinosum, cysteine synthase was found to be constitutive, while in the Rhodospirillaceae tested the enzyme is repressed by sulfide. Thiosulfate had a derepressive effect in Rhodopseudomonas globiformis but strongly repressed cysteine synthase in R. sulfidophila and R. palustris. Cysteine had only moderate effects with the species tested.     

Transformation of metals and metal ions by hydrogenases from phototrophic bacteria

The ability of hydrogenases isolated from Thiocapsa roseopersicina and Lamprobacter modestohalophilus to reduce metal ions and oxidize metals has been studied. Hydrogenases from both phototrophic bacteria oxidized metallic Fe, Cd, Zn and Ni into their ionic forms with simultaneous evolution of molecular hydrogen. The metal oxidation rate decreased in the series Zn>Fe>Cd>Ni and depended on the pH. The presence of methyl viologen in the reaction system accelerated this process. T. roseopersicina and L. modestohalophilus cells and their hydrogenases reduced Ni(II), Pt(IV), Pd(II) or Ru(III) to their metallic forms under H₂ atmosphere. These results suggest that metals or metal ions can serve as electron donors or acceptors for hydrogenases from phototrophic bacteria.     

Formation of poly(3-hydroxyalkanoates) by phototrophic and chemolithotrophic bacteria

The formation of poly(3-hydroxyalkanoic acid), PHA, by various strains of chemolithotrophic and phototrophic bacteria has been examined. Chemolithotrophic bacteria were grown aerobically under nitrogen-limiting conditions on various aliphatic organic acids. Phototrophic bacteria were grown anaerobically in the light on a nitrogen-rich medium and were subsequently transferred to a nitrogen-free medium containing acetate, propionate, valerate, heptanoate or octanoate as carbon source. All 41 strains investigated in this study were able to synthesize and accumulate PHA. All 11 strains of chemolithotrophic bacteria and all 15 strains belonging to the non-sulfur purple bacteria synthesized a polymer, which contained 3-hydroxy-valerate (3HV) beside 3-hydroxybutyrate (3HB), if the cells were cultivated in the presence of propionate, valerate or heptanoate. Many non-sulfur purple bacteria synthesized copolyesters of 3HB and 3HV even with acetate as carbon source. In contrast, most sulfur purple bacteria did not incorporate 3HV at all. Among 15 strains tested, only Chromatium vinosum strain 1611, C. purpuratum strain BN5500 and Lamprocystis roseopersicina strain 3112 were able to synthesize polyesters containing 3HV with propionate, valerate or heptanoate as carbon source.     

Identification of phototrophic sulfur bacteria through the analysis of lmwRNA band patterns

Several phototrophic sulfur bacteria were identified preliminarily through the analysis of the low-molecular-weight RNA fraction (lmwRNA) of bacterial cells. This fraction includes the ribosomal 5S RNA and several transfer RNAs. These molecules were separated by high-resolution electrophoresis in polyacrylamide gels, and the resulting band patterns were used as fingerprints for the identification of the organisms. We examined a large number of well-characterized reference strains together with a broad range of purple sulfur bacterial isolates from freshwater and marine environments. A cluster analysis was run using the similarity matrix calculated from the band patterns. Despite the shortcomings of the method, close relatives were clustered together yielding a number of groups consistent with the phylogenetic arrangement established through the analyses of a few available 16S rRNA gene sequences. Thus, the classification obtained gives further support to rearrangement of the group as the analyses of 16S rRNA gene sequences had previously suggested. We conclude that the analysis of lmwRNA band patterns is a rapid and simple tool for grouping and preliminarily identifying new isolates of phototrophic sulfur bacteria. Received: 5 February 1998 / Accepted: 15 June 1998     

Noncyclic electron transport in chromatophores from photolithotrophically grown Rhodobacter sulfidophilus

Chromatophores isolated from the marine phototrophic bacterium Rhodobacter sulfidophilus were found to photoreduce NAD with sulfide as the electron donor. The apparent K _m for sulfide was 370 M and the optimal pH was 7.0. The rate of NAD photoreduction in chromatophore suspensions with sulfide as the electron donor (about 7–12 M/h·mol Bchl) was approximately onetenth the rate of sulfide oxidation in whole cell suspensions. NAD photoreduction was inhibited by rotenone, carbonyl cyanide-m-chlorophenylhydrazone, and antimycin A. Sulfide reduced ubiquinone in the dark when added to anaerobic chromatophore suspensions. These results suggest that electron transport from sulfide to NAD involves an initial dark reduction of ubiquinone followed by reverse electron transport from ubiquinol to NAD mediated by NADH dehydrogenase.Abbreviations Bchl bacteriochlorophyll - CCCP carbonyl cyanide-m-chlorophenylhydrazone - MOPS 3(N-morpholino)-propane sulfonate - Uq ubiquinone     

Degradation of p-nitrophenol by the phototrophic bacterium Rhodobacter capsulatus

The phototrophic bacterium Rhodobacter capsulatus detoxified p-nitrophenol and 4-nitrocatechol. The bacterium tolerated moderate concentrations of p-nitrophenol (up to 0.5 mM) and degraded it under light at an optimal O₂ pressure of 20 kPa. The bacterium did not metabolize the xenobiotic in the dark or under strictly anoxic conditions or high O₂ pressure. Bacterial growth with acetate in the presence of p-nitrophenol took place with the simultaneous release of nonstoichiometric amounts of 4-nitrocatechol, which can also be degraded by the bacterium. Crude extracts from R. capsulatus produced 4-nitrocatechol from p-nitrophenol upon the addition of NAD(P)H, although at a very low rate. A constitutive catechol 1,2-dioxygenase activity yielding cis,cis-muconate was also detected in crude extracts of R. capsulatus. Further degradation of 4-nitrocatechol included both nitrite- and CO₂-releasing steps since: (1) a strain of R. capsulatus (B10) unable to assimilate nitrate and nitrite released nitrite into the medium when grown with p-nitrophenol or 4-nitrocatechol, and the nitrite concentration was stoichiometric with the 4-nitrocatechol degraded, and (2) cultures of R. capsulatus growing microaerobically produced low amounts of ¹⁴CO₂ from radiolabeled p-nitrophenol. The radioactivity was also incorporated into cellular compounds from cells grown with uniformly labeled ¹⁴C-p-nitrophenol. From these results we concluded that the xenobiotic is used as a carbon source by R. capsulatus, but that only the strain able to assimilate nitrite (E1F1) can use p-nitrophenol as a nitrogen source. Received: 30 December 1996 / Accepted: 3 September 1997     

Sedimentary photosynthetic pigments of algae and phototrophic bacteria in Lake Hamana,Japan: temporal changes of anoxia in its five basins

We analyzed photosynthetic pigments of algae and bacteria (phototrophic sulfur bacteria: Chromatium and brown Chlorobium) in sediment cores and water samples obtained from five basins of Lake Hamana, a brackish, eutrophic, holomictic lake in Japan, and discussed our findings in relation to the distribution of the phototrophs. The four outer basins are connected to the central basin by narrow inlets. The prevalence of anoxia in Lake Hamana was demonstrated by the widespread presence of bacterial pigments in each core. The construction of training walls in 1954–1956 to direct tidal currents into the lake via Imagire-guchi Channel, the sole inlet for seawater, increased the lake water circulation, suppressed the development of anoxia, and caused Chromatium to disappear. Strong correlations (r ² 0.7) between total algal carotenoid (TAC) and total bacterial carotenoid (TBC) contents in each core were found in four basins. We ascribe this to the induction of anoxia by water stratification and algal proliferation, which precede the growth of phototrophic sulfur bacteria in the deeper layers of the water column. The slopes of the TBC–TAC correlations in the sediment cores, indicating the extent and stability of anoxia at each site, differed among basins (0.23–0.67) and were inversely related to the exchange rate of water by seawater intrusion in each basin.     

The sulfide affinity of phototrophic bacteria in relation to the location of elemental sulfur

Seventeen strains of phototrophic bacteria (4 strains of Chromatium spp., 2 strains of Thiocapsa sp., 4 strains of Ectothiorhodospira spp., 2 strains of Rhodopseudomonas sp., and 5 strains of Chlorobium spp.) have been grown in sulfide-limited continuous cultures to assess the affinity for sulfide. It was found that the affinity (calculated as the initial slope of the specific growth rate versus the concentration of sulfide) is higher in those phototrophic bacteria that deposit elemental sulfur outside the cells, than in those bacteria that store the sulfur inside the cells. A hypothesis is presented to explain this correlation.Dedicated to Prof. Dr. Hans G. Schlegel on the occasion of his 60th birthday     

Simultaneous phototrophic and chemotrophic growth in the purple sulfur bacterium Thiocapsa roseopersicina M1

Abstract The anoxygenic phototrophic purple sulfur bacterium Thiocapsa roseopersicina was grown in illuminated continuous cultures with thiosulfate as growth limiting substrate. Aeration resulted in completely colorless cells growing chemotrophically, whereafter the conditions were changed to a 23 h oxic/1 h anoxic regime. After 11 volume changes at a dilution rate of 0.031 h⁻¹ (35% of μ_max) a time dependent equilibrium was established. During the 23 h oxic periods bacteriochlorophyll a synthesis (BChl a ) was not observed, whereas during the 1 h anoxic periods synthesis was maximal (i.e. 1.1 μg (mg protein)⁻¹ h⁻¹). As a result the BChl a concentration gradually increased from zero to an average value over 24 h of 1.9 μg (mg protein)⁻¹. Concomitantly, the protein concentration increased from 13.9 mg 1⁻¹ during continuous oxic conditions to 28.8 mg 1⁻¹. For comparison, the protein concentration during fully phototrophic growth at an identical thiosulfate concentration in the inflowing medium was 53.7 mg 1⁻¹. The specific respiration rate was 8 μmol O₂ (mg protein)⁻¹ h⁻¹ during full chemotrophic growth and gradually decreased to 3.5 μmol O₂ (mg protein)⁻¹ h⁻¹ after 11 volume changes at the regime employed. These data show that T. rosepersicina is able to simultaneously utilize light and aerobic respiration of thiosulfate as sources of energy. The ecological relevance of the data is discussed.     

Quinones of phototrophic purple bacteria

Abstract The quinone composition of the recognized species of the phototrophic purple nonsulfur bacteria, the Ectothiorhodospiraceae, and some Chromatiaceae species has been determined. Altogether more than 50 strains of 33 species have been investigated. Some of the purple nonsulfur bacteria have Q-10 as sole quinone component, while others have Q-10, Q-9, or Q-8, respectively, together with menaquinones of the same isoprenoid chain length as the major components. Rhodoquinone is present in Rhodospirillum rubrum and Rhodospirillum photometricum . The Ectothiorhodospira species have either Q-8 and MK-8, like the Chromatiaceae species, or Q-7 and MK-7 as the major components.     

青海湖裸鲤肠道乳酸菌多样性与抑菌活性

【目的】通过生理生化特性,结合16S r RNA基因序列分析研究青海湖裸鲤肠道乳酸菌分离株的多样性,并对这些代表株的抑菌活性进行初步探讨,以期筛选具有高效抑菌活性的鱼源益生菌。【方法】对分离的47株乳酸菌代表株进行p H、温度生长范围、耐盐性等生理生化特征检测,结合16S r RNA基因序列对已分离到的乳酸菌进行基因分型和菌种鉴定,采用牛津杯双层平板法检测乳酸菌代表株的抑菌活性。【结果】鉴定结果显示:23株为Lactobacillus fuchuensis(48.94%),12株为Lactobacillus curvatus(25.53%),3株为Leuconostoc fallax(6.38%),2株为Lactobacillus sakei(4.26%),2株为Weissella ceti(4.26%);2株为Lactococcus cremoris(4.26%),1株为Leuconostoc lactis(2.13%),1株为Weissella minor(2.13%),1株为Enterococcus devriesei(2.13%)。qz1217、qz1196、qz1220所在的A、B、C三组乳酸菌在5-50°C的温度范围内生长良好,qz1196、qz1220所在的B、C组在pH 3.0-10.0的范围内生长良好,几乎所有乳酸菌都具有耐6.5%盐浓度特性。13株乳酸菌菌株对6种病原菌都具有抑制作用。通过排除酸、过氧化氢实验,发现上清液仍然具有抑菌活性。对qz1251发酵液进行蛋白酶处理,抑菌活性消失,确定其抑菌物质属于蛋白类物质,是一种细菌素。【结论】青海湖裸鲤肠道附着乳酸菌的多样性为益生性乳酸菌的筛选提供优质资源及数据参考。     

Dechlorination of 2,3,5,6-tetrachlorobiphenyl by a phototrophic enrichment culture

A phototrophic enrichment culture, using acetate as carbon source, reductively dechlorinated 2,3,5,6-tetrachlorobiphenyl. ortho chlorines were removed preferentially over meta chlorines. Tri- and dichlorobiphenyls were the major products. During 14 months incubation, chlorine was removed from 58% of the target molecules; 19% of the total chlorines were removed. Dechlorination did not occur in a control culture incubated in the dark.     

Isolation and growth of the phototrophic bacterium Rhodopseudomonas palustris strain B1 in sago-starch-processing wastewater

An indigenous strain of the purple non-sulphur phototrophic bacterium, Rhodopseudomonas palustris strain B1, was selected for the utilization and treatment of wastewater from a sago-starch-processing decanter. Growth of Strain B1 under anaerobic–light conditions in the carbohydrate-rich effluent was optimized by using 50% (v/v) effluent diluted in a basal minimal mineral medium with the addition to 0.1% (w/v) yeast extract. The optimum level of nitrogen source supplement, ammonium sulphate, was 1.0g/l. Highest cell mass concentration was achieved by using tungsten lamps as the light source with a light intensity of 4 klux. Under these optimal conditions, a maximum biomass of about 2.5g dry cell/l with a pigment content of about 1.1mg carotenoid/g dry weight cell was achieved after 96h of anaerobic cultivation. There was a 77% reduc n the chemical oxygen demand (COD) of the effluent. A cell yield of about 0.59g dry weight cell/g COD was obtained.     

Expression and characterization of the assimilatory NADH-nitrite reductase from the phototrophic bacterium Rhodobacter capsulatus E1F1

A nas gene region from Rhodobacter capsulatus E1F1 containing the putative nasB gene for nitrite reductase was previously cloned. The recombinant His₆-NasB protein overproduced in E. coli showed nitrite reductase activity in vitro with both reduced methyl viologen and NADH as electron donors. The apparent K _m values for nitrite and NADH were 0.5 mM and 20 μM, respectively, at the pH and temperature optima (pH 9 and 30°C). The optical spectrum showed features that indicate the presence of FAD, iron-sulfur cluster and siroheme as prosthetic groups, and nitrite reductase activity was inhibited by sulfide and iron reagents. These results indicate that the phototrophic bacterium R. capsulatus E1F1 possesses an assimilatory NADH-nitrite reductase similar to that described in non-phototrophic organisms.     

Purification and properties of phototrophic bacteria Thiocapsa roseopersicina hydrogenase bound with chromatophores]

The method of solution and puridication of hydrogenase from chromatophores of purpur sulphur bacteria Thiocapsa roseopersicina strain BBS are described. Hydrogenase molecular weight is 73000. It contains 4,4 mole S2- and 3.1 mole Fe2+ per mole of protein; pI 4.15. The enzyme absorption spectrum has the maximun et 400-410 nm, which is characteristic of proteins containing non-haem iron. Membrane--linked enzyme as well as soluble hydrogenase of that microorganism is characterized by high thermal stability: inactivation occurs at the temperature above 78 degrees C when the optimal temperature for that enzyme is 70 degrees C. Homogenous enzyme catalyses D2--H2O exchange reaction, reversible redox reaction of methyl viologene and benzyl viologene.