Green Fluorescent Pigment Research Articles

Macro- and micro-scale adaptations allow distinct Stylophora pistillata-symbiodiniaceae holobionts to optimize performance across a broad light habitat.

In sessile organisms, phenotypic plasticity represents an important strategy for dealing with environmental variability. Here we test if phenotypic plasticity enables the common coral Stylophora pistillata to occupy a broad niche. We find clear differences in the photo-physiology of four putative species of photosynthetic dinoflagellate symbionts associated with the coral S. pistillata, namely, Cladocopium 'C35a', 'C79', 'C78a' and 'C8a'. Coral phenotypic responses were also tightly linked to symbiont identity. Corals with Cladocopium 'C8a' have more "open" macro-morphology compared to colonies associating with depth-restricted Cladocopium 'C35a' or 'C78a' in the same shallow water habitat. Corals with Cladocopium 'C8a' had 40 to 60% lower symbiont cell densities compared to other holobionts but were more efficient at acclimating over a range of light levels, with clear mechanisms to dissipate excess light energy. This holobiont contains host-based green fluorescent pigments, increased concentrations of symbiont-based mycosporine amino acids, and xanthophyll cycling in high light habitats. Photosynthetic efficiency was also adjusted over the light habitat. In contrast, limited micro-scale responses were observed between three depth-restricted symbionts: Cladocopium 'C79', 'C35a', and 'C78a'. To optimize light levels reaching the photosynthetic unit, these colonies rely on a more closed macro-morphology under high light levels, which reduces incident light levels by up to 43%, and higher symbiont densities . Our results show that distinct macro- and micro-scale adaptations lead to functional differences between four distinct S. pistillata holobionts, allowing them to co-exist by filling specific niches on a small, but environmentally diverse, spatial scale. Key index words: Light, Symbiodiniaceae, coral, pigments, Stylophora pistillata, ITS2, phenotypic plasticity, niche diversification.

ISOLATION AND IDENTIFICATION OF ANTIBIOTIC PRODUCING PSEUDOMONAS FLUORESCENS NBRC-14160 FROM DELTA SOIL IN EGYPT

Pseudomonas fluorescens is one of plant Growth Promoting Rhizobacteria (PGPR) members which has a major role in the biological control of bacterial and fungal pathogens. A research was conducted at the Laboratory of Microbiology, Faculty of Agriculture, Ain Shams University, Qalubyia governorate intended for isolating Pseudomonas fluorescens isolate, efficient in antibiotic production. For isolation, soil samples, collected from Faculty of agriculture farm soil at depth of 10 cm, were screened for Ps. fluorescens isolates by cultivating the samples on King’s medium. Out of 30 isolates obtained, one was selected based on its high lipid content, because of the lipid’s correlation to antibiotic production and inhibitory activity. The selected isolate was characterized by morphological, physiological, biochemical tests then confirmed its identity by 16S rRNA gene sequence analysis and named Pseudomonas fluorescens NBRC-14160 16srRNA. Morphological features showed that Ps. fluorescens NBRC-14160 has large colonies, with irregular surface, opaque, producing green fluorescent pigments. Cells are short rods, Gram negative. Physiological features indicated that Ps. fluorescens NBRC-14160 is capable of producing several exoenzymes including lipase, phospholipase, protease and chitinase and incapable of producing amylase and cellulase. Carbohydrate fermentation tests were positive for fructose, glucose, D-glucose, and galactose, forming acids after 24 hrs. of incubation at 30˚C. However, it loses the ability to ferment inositol, mannose, xylose, mannitol, raffinose and rhamnose sugars. It’s lipid content was 607 mg/g. Inhibitory activity was studied by the method of disc diffusion test against nine pathogenic bacterial and fungal strains. Staphylococcus aureus was the most sensitive bacterial pathogen towards P. fluorescens NBRC-14160 with a 4.5 cm zone of inhibition, while Serratia marscens had 0.95 cm zone. Aspergillus niger and Alternaria solani were the most sensitive fungal pathogens towards P. fluorescens NBRC-14160 with 2.55 and 2.5 cm of inhibition zone, respectively. However, the most resistant fungus was Fusarium oxysporum with 1.3 cm inhibition zone.

Isolation and identification of antibiotic producing Pseudomonas fluorescens NBRC-14160 from Delta Soil in Egypt

Pseudomonas fluorescens is one of plant Growth Promoting Rhizobacteria (PGPR) members which has a major role in the biological control of bacterial and fungal pathogens. A research was conducted at the Laboratory of Microbiology, Faculty of Agriculture, Ain Shams University, Qalubyia governorate intended for isolating Pseudomonas fluorescens isolate, efficient in antibiotic production. For isolation, soil samples, collected from Faculty of agriculture farm soil at depth of 10 cm, were screened for Ps. fluorescens isolates by cultivating the samples on King’s medium. Out of 30 isolates obtained, one was selected based on its high lipid content, because of the lipid’s correlation to antibiotic production and inhibitory activity. The selected isolate was characterized by morphological, physiological, biochemical tests then confirmed its identity by 16S rRNA gene sequence analysis and named Pseudomonas fluorescens NBRC-14160 16srRNA. Morphological features showed that Ps. fluorescens NBRC-14160 has large colonies, with irregular surface, opaque, producing green fluorescent pigments. Cells are short rods, Gram negative. Physiological features indicated that Ps. fluorescens NBRC-14160 is capable of producing several exo-enzymes including lipase, phospholipase, protease and chitinase and incapable of producing amylase and cellulase. Carbohydrate fermentation tests were positive for fructose, glucose, D-glucose, and galactose, forming acids after 24 hrs. of incubation at 30˚C. However, it loses the ability to ferment inositol, mannose, xylose, mannitol, raffinose and rhamnose sugars. It’s lipid content was 607 mg/g. Inhibitory activity was studied by the method of disc diffusion test against nine pathogenic bacterial and fungal strains. Staphylococcus aureus was the most sensitive bacterial pathogen towards P. fluorescens NBRC-14160 with a 4.5 cm zone of inhibition, while Serratia marscens had 0.95 cm zone. Aspergillus niger and Alternaria solani were the most sensitive fungal pathogens towards P. fluorescens NBRC-14160 with 2.55 and 2.5 cm of inhibition zone, respectively. However, the most resistant fungus was Fusarium oxysporum with 1.3 cm inhibition zone.

Study of isolation method of green fluorescent pigments contained in cherry tomatoes using column chromatography

We investigated a method to isolate the green fluorescent pigments contained in cherry tomatoes using column chromatography. To determine the best developing solvent for the column chromatography, we tested several solvents using a thin-layer chromatography and adopted a solution containing hexane and 2-propanol in a ratio of 99:1 because its Rf value can be adjusted to 0.3. Some pigments were extracted by the column chromatography and identified by evaluating their absorption spectra: β-carotene, phytoene, phytofluene, lycopene and ζ-carotene. Among them, we observed photoluminescence emissions from the solutions containing phytofluene and lycopene. In particular, lycopene reveals a green emission around 542 nm. However, the solution containing lycopene reveals another undesired red emission around 674 nm that originates from the chlorophylls. By performing the second column chromatography with a different developing solvent, only lycopene was successfully isolated to produce a pure green emission.

First Report of Pseudomonas syringae pv. syringae Associated with Bacterial Blossom Blast on Apple (Malus pumila) in the United States

In May 2017, dead buds at pink and tight cluster growth stages were observed on young apple trees of cultivar SnapDragon at two orchard sites in Marlboro, NY. Disease symptoms included necrosis of lateral and terminal flower buds that were wilted and drooping. Bark below the dead buds on severely affected trees showed black streaking with or without interspersing white streaks. On the cross-section of branch wood, just below the buds, light brown xylem and glossy water-soaked sections resembling frost damage were observed. After bark shaving, dark xylem tissue was visible along the branch length, in contrast to the light or pale color of the healthy tissue. The incidence of the disease on trees was 3 to 5%, and 0.5% of trees died. There were five near-frost events recorded on 4, 8, 9, 10, and 11 May 2017, leading us to suspect that the observed symptoms were blossom blast caused by the ice-nucleating strains of Pseudomonas syringae. Isolation from the affected tree samples was performed on King’s B (KB) and nutrient sucrose agar (NSA) media. After 3 days at 25°C, most of bacterial isolates formed round, shiny, gray-white colonies producing green fluorescent pigment on KB medium and typical levan-positive colonies on NSA. Eight isolates, inducing a hypersensitive reaction on tobacco plants, were further characterized by conventional bacteriological and molecular tests (Braun-Kiewnick and Sands 2001). The analyzed bacterial isolates were Gram-negative, aerobic, hydrolyzed gelatin and aesculin, but not pectate, and were negative for oxidase and arginine-dihydrolase activity. All strains were ice-nucleation active and utilized sucrose, sorbitol, and inositol as carbon sources, but not tyrosine and tartaric acid. Results of levan, cytochrome c oxidase, potato soft rot, arginine dihydrolase, and tobacco hypersensitive reaction (LOPAT) tests indicated that the isolates belong to P. syringae group Ia, and gelatin hydrolysis, aesculin hydrolysis, tyrosinase activity, and tartrate utilization (GATTa) tests designated them as pathovar (pv.) syringae (Kaluzna et al. 2012). Polymerase chain reaction (PCR) with specific primers B1/B2 (Sorensen et al. 1998) and SyD1/SyD2 (Bultreys and Gheysen 1999) showed that isolates possessed genes responsible for synthesis (syrB) and secretion (syrD) of syringomycin, respectively. Partial sequencing of the 16S rDNA gene of three isolates (GenBank MG788015 to MG788017) showed 99 to 100% identity with sequences of P. syringae pv. syringae previously deposited in the NCBI GenBank database. To complete Koch’s postulates, pathogenicity of the four isolates was confirmed by inoculation of 1-month-old potted apple plantlets grown from seeds collected from mature apple fruit of cultivar SnapDragon. One plant per isolate was inoculated multiple times as follows: using a needleless syringe, two leaves per plant were spot-infiltrated with bacterial suspension (10⁶ CFU/ml); on the same plants, stem was also inoculated by inserting a sterile toothpick previously dipped into a bacterial colony. Sterile distilled water was used as a negative control. Inoculated plantlets were sealed within plastic bags for 48 h and further incubated at 22°C. Inoculation test was repeated in two independent experiments. Necrosis of the leaf and stem tissue developed at the point of inoculation, and its progression was observed daily for 2 weeks. No symptoms developed on the control plants. Bacteria were reisolated from inoculated tissues and confirmed as P. syringae pv. syringae by PCR for syrB and syrD genes. This is the first report of P. syringae pv. syringae causing blossom blast symptoms on developing apple buds in commercial orchards in the United States. Even though it rarely occurs on apples, with most recent frosts significantly lowering apple yields in Michigan (2012) and New York (2016), blossom blast might become a more common problem in U.S. orchards. Previously, P. syringae pv. syringae was isolated from dormant apple buds without and with internal necrosis in the United States (Burr and Katz 1984).

Pseudomonas viridiflava in imported carrot seeds

In 2005, a sample of carrot seeds var. Kikuyo imported from Japan was submitted to the Instituto Biológico for bacteriological phytosanitary certification. Circular, convex and creamy colored colonies were obtained. These isolates produced a green fluorescent pigment on King’s B medium and incited a hypersensitivity reaction in tobacco leaves. They were oxidase and arginine dihydrolase negative, did not produce levan but caused soft rot in potato slices and were identified as Pseudomonas viridiflava. Artificial inoculations by infiltration of bacterial suspensions at the crown area or by placing a drop of bacterial suspension on this area and pricking with a sterile needle, produced light to dark brown areas around the injured points 5 days after inoculation. The bacterium was then re-isolated. Although this bacterium could be an opportunistic pathogen and was isolated from asymptomatic samples, eventual losses in the field and post-harvest storage is possible.

First Report of Bacterial Leaf Spot of Basil Caused by Pseudomonas viridiflava in Hungary.

In April 2011, typical bacterial spot symptoms were observed on sweet basil plantlets (Ocimum basilicum L.) in a supermarket in Budapest, Hungary. Affected plants had dark brown-to-black lesions on the cotyledons. Spots on the leaves were first water soaked and then became necrotic and progressed inward from the margins. Symptoms were similar to those reported by Little et al. (3) on basil affected by Pseudomonas viridiflava. Bacteria consistently isolated from leaf lesions formed mucoid colonies with a green fluorescent pigment on King's B medium. Strains were gram negative. In LOPAT (levan-oxidase-potato rot-arginine dihydrolase-tobacco hypersensitivity) tests (2), all induced a hypersensitive reaction (HR) in tobacco (Nicotiana tabacum L. cv. White Burley) leaves (1), caused soft rot of potato tuber slices, and were negative for levan, oxidase, and arginine dihydrolase. Biochemical tests, API 20NE and API 50 CH (Biomérieux, Marcy l'Etoile, France), were also used for identification. The pathogenicity of three isolates was tested twice by injecting 20-day-old healthy basil plants with a bacterial suspension (107 CFU/ml). Controls were injected with sterile distilled water. Plants were kept at 25 to 28°C and 80 to 100% relative humidity. Forty-eight hours after inoculation, dark brown-to-black lesions were observed only on inoculated plants. The bacterium was reisolated from lesions of all plants tested, fulfilling Koch's postulates. No lesions were observed on controls. To identify the pathogen, a PCR technique was used. The 16SrDNA region was amplified with general bacterial primer pair (63f forward and 1389r reverse) (4) then the PCR products were cloned into Escherichia coli DH5α cells and a recombinant plasmid was sequenced by M13 forward and reverse primers. The sequence was deposited in GenBank (Accession No. HE585219). On the basis of the symptoms, biochemical tests, and 16SrDNA sequence homology (99% sequence similarity with a number of P. viridiflava isolates), the pathogen was identified as P. viridiflava. To our knowledge, this is the first report of bacterial leaf spot of basil in Hungary, which can seriously affect the basil production.

Towards Raman‐based epidemiological typing of Pseudomonas aeruginosa

Raman spectra of bacteria can be used as highly specific fingerprints, enabling discrimination at strain level. Pseudomonas aeruginosa strains can be strongly pigmented, making it difficult to obtain high quality spectra of such isolates due to high fluorescent spectral backgrounds. Furthermore, the spectra that could be measured with acceptable quality often showed large spectral variations limiting the reproducibility required for strain level discrimination. P. aeruginosa produces a characteristic yellowish green fluorescent pigment, called pyoverdin. Applying a washing procedure to reduce the amount of fluorescent pigment, enabled the highly pigmented isolates to be measured with sufficient spectral quality. Isolation of the pigment/pyoverdin spectral features, together with spectral scaling methods improved reproducibility. It will be important to analyze the range of the spectral variations that can occur and ensure the correction of all of these factors to obtain the highest reproducibility required for strain level typing.

Pseudomonas brassicacearum subsp. neoaurantiaca subsp. nov., orange-pigmented bacteria isolated from soil and the rhizosphere of agricultural plants

A large group of 38 strains of saprophytic bacteria was isolated from soil and the rhizosphere of agricultural plants. The novel organisms were Gram-negative, aerobic, rod-shaped bacteria that produced a green fluorescent pigment, a red-orange diffusible pigment and a complex mixture of phloroglucinol derivates with antimicrobial activity. The latter have not been found in other bacteria, but are peculiar to ferns. The bacteria were vigorous denitrifiers that synthesized levan from sucrose and liquefied gelatin, but were found not to degrade aesculin, starch, agar, Tween 80 or DNA. Bacterial growth was found to occur at 4 degrees C but not at 40 degrees C. The predominant cellular fatty acids were 16 : 0, 16 : 1(n-7), 18 : 1(n-7) and 17 : 0 cyclo. The G+C content of the novel bacteria was 61.0-62.9 mol%. 16S rRNA gene sequence analysis indicated that the representative strain CIP 109457(T) had a clear affiliation with Pseudomonas sensu stricto groups, with the nearest relatives being Pseudomonas brassicacearum, P. thivervalensis, P. corrugata, P. mediterranea and P. kilonensis. DNA-DNA hybridization experiments showed that the group of isolated strains exhibited high levels of genetic relatedness (81-100 %), confirming that they are representatives of the same species. At the same time, they bound at low levels (4-46 %) with DNA of the type strains of their nearest relatives with the exception of P. brassicacearum; DNA binding of 90 % with the DNA of P. brassicacearum CIP 107059(T) suggested that the bacteria studied belong to this species. Analysis of taxonomic data indicated that the group of novel bacteria maintain a distinct phenotypic profile, allowing the description of novel subspecies within P. brassicacearum, for which the following names are proposed: Pseudomonas brassicacearum subsp. brassicacearum subsp. nov. (type strain DBK11(T) =CFBP 11706(T) =CIP 107059(T) =DSM 13227(T) =JCM 11938(T)) and Pseudomonas brassicacearum subsp. neoaurantiaca subsp. nov., with the type strain CIP 109457(T) (=ATCC 49054(T) =IMV 387(T) =VKM B-1524(T)).

First Report of Bacterial Leaf Spot of Basil Caused by Pseudomonas viridiflava in Argentina.

In June 1998, during a cool, humid period, typical bacterial spot symptoms were observed on basil plantlets (Ocimun basilicum L. 'Royal Louis' and 'Zaes') in a commercial greenhouse in La Plata, Argentina. Affected plants had dark brown to black lesions on cotyledons. Spots on leaves were first water soaked, then became necrotic and progressed inward from the margins. Disease incidence approached 30%. Symptoms were similar to those reported by Little et al. (2) on basil affected by Pseudomonas viridiflava. No pathogenic fungi or viruses were associated with symptomatic plants. Bacterial streaming was observed from lesion margins. Bacteria consistently isolated from leaf lesions formed cream-colored, glistening, convex colonies on sucrose peptone agar and a green fluorescent pigment on King's medium B. Bacterial growth produced a distinctive olive green pigment on glycerol agar medium and a pink pigment on T-5 medium (1). Four isolates selected for further study were aerobic, Gram-negative, non-spore-forming rods. In LOPAT (levan-oxidase-potato rot-arginine dihydrolase-tobacco hypersensitivity) tests, all induced a hypersensitive response in tobacco plants, caused soft rot of potato tubers, and were negative for levan, oxidase, and arginine dihydrolase. In addition, strains rotted onion slices and produced a reddish sunken lesion on bean pods. Acid was produced aerobically from D-glucose, mannitol, mesoinositol and sorbitol, but not from D-arabinose, L-rhamnose, melibiose, amygdalin, or sucrose. Bacteria used D-tartrate, pyruvate, and citrate, but not benzoate. The strains did not hydrolyze starch, exhibited an oxidative metabolism of glucose, and did not reduce nitrates to nitrites or accumulate poly-β-hydroxybutyrate inclusions. Negative reactions were obtained with indole, ornithine, and D-tryptophan. Isolates hydrolyzed gelatine, used Tween 80, were positive for catalase, and were unable to grow in the presence of 5% NaCl. Colonies developed at 4°C but not 37°C. Reactions were identical to those of reference strains ICMP 5776 and 12363, which were included in all tests for comparison. Pathogenicity was verified on 35-day-old basil plants by both spraying and infiltration inoculations with bacterial suspensions (108 and 105 cells per ml, respectively). Carborundum was included in the inoculum used for a set of plants inoculated by spraying. Controls were injected or sprayed (with and without Carborundum) with sterile, distilled water. In addition, bean (Phaseolus vulgaris cv. Nag12 INTA) and lettuce (Lactuca sativa cv. criolla), both reported as host plants, were inoculated by spraying with bacterial suspensions of 107 cells per ml plus Carborundum. After 48 h in a humid chamber, inoculated plants and controls were maintained at 23 ± 3°C. Symptoms on basil plants inoculated by injection or spraying with Carborundum were identical to those observed on basil in the field. Symptoms on bean and lettuce were similar to those described for P. viridiflava. The bacterium was reisolated from lesions of all species tested, fulfilling Koch's postulates. No lesions were observed on controls or on plants sprayed without Carborundum, suggesting that bacteria gain entry through wounds. The microorganism was identified by physiological tests and polymerase chain reaction as P. viridiflava. This is the first report of bacterial leaf spot of basil in Argentina.

Pseudomonas fuscovaginae sp. nov., nom. rev.

The name Pseudomonas fuscovaginae Tanii, Miyajima and Akita 1976 was omitted from the Approved Lists of Bacterial Names. Therefore, this name is here revived for the organism to which it originally referred. The cells of strains of this species are aerobic, gram negative, and rod shaped with polar flagella. They oxidize glucose in oxidation-fermentation medium, and they produce a green fluorescent pigment, oxidase, and arginine dihydrolase. Denitrification, β-glucosidase, pit formation on polypectate gel and growth at 37°C are negative. Characteristics that distinguish this species from other fluorescent pseudomonads which are positive for arginine dihydrolase and oxidase are its ability to produce a hypersensitivity reaction in tobacco plants and its inability to utilize 2-ketogluconate or inositol. These bacteria were pathogenic to Oryza sativa and eight other species of the Gramineae. The type strain is NCPPB 3085 (= PDDCC 5940).

Ageing of Neurospora crassa. I. Evidence for the free radical theory of ageing from studies of a natural-death mutant

A recessive mutant of Neurospora crassa, called natural-death, is characterized by a decreasing clonal growth potential under all nutritional conditions and the irreversible cessation of growth. The primary molecular defect of this mutant is not known. Evidence presented here, based upon measurements of the activities and thermolabilities of several enzymes, suggests that faulty protein synthesis is probably not a cause of the senescence and death of the mutant, as suggested by Lewis and Holliday ( Nature, 228 (1970) 877). Three lines of evidence indicate that lipid autoxidation and associated free radical reactions contribute to the senescence and death of this mutant: (1) The relative times before the onset of senescence and death of mutant clones in the last 40% of their chronological life-span were prolonged 2 to 3-fold by either dietary antioxidants or selenite and the total life-span was increased by 40% to 80%. These compounds also alleviated the senescent morphology and enhanced biomass production; (2) Senescing clones accumulated a green fluorescent pigment in situ, but dietary antioxidant nordihydroguaiaretic acid prevented this accumulation. The fluorescent pigment exhibited the spectral properties of lipofuscin, an end product of lipid autoxidation; (3) Relative to wild type, mycelial extracts of the mutant exhibited a 2 to 4-fold excess of activities of the antioxygenic enzymes superoxide dismutase, glutathione peroxidase and glutatione reductase. We briefly review: (1) the roles of antioxygenic enzymes and antioxidants in their protection against cellular damage from lipid autoxidation and free radical reactions; and (2) the major lines of evidence which appear to support a form of the free radical theory of ageing, encompassing the interrelated processes of membrane deterioration, lipid autoxidation and deleterious free radical reactions as the major causes of cellular deterioration.

Influence of Temperature on Some Biochemical Characteristics of Pseudomonas Associated with Spoilage of Chicken

SUMMARY Studies were conducted with four cultures of Pseudomonas isolated from frozen chicken. The effect of temperature on some biochemical activities of the organisms was evaluated and the individual response of the cultures to temperature was determined. Growth, survival and production of the green fluorescent pigment, pyoverdine, and extra‐cellular proteinase and lipase activities were used as indices of the ability of pseudomonads to produce spoilage. The four isolates differed in their ability to perform the metabolic functions mentioned. The cultures were incubated at 15°, 5°, −18°, and −29°C. Assays for proteolysis were made by means of a dye binding method; lipolysis was determined by titration of free fatty acids released from chicken fat, and a photo‐fluorometer was used to measure fluorescent pigment. Growth was determined by colony count. At temperatures above 0°C, survival was better and growth and enzyme activity were more extensive at 5° than at 15°C. Proteinase activity increased continuously, even when viable cells were decreasing; lipase production was correlated with growth. Formation of pyoverdine declined faster than did cell numbers. Survival of the cultures was better at −18° than at −29°C, Impairment of pyoverdine secretion was observed after exposure of the organisms to freezing temperatures, but the activity of the extracellular enzymes was not affected at temperatures below 0°C. No marked differences were observed among cultures in rate of cell division, but maximum populations, survival of organisms and stability of the proteolytic, lipolytic and fluorescent activities of the isolates were inversely related to biochemical activity above 0°C.

Clover rhizoplane bacteria antagonistic to Rhizobium trifolii.

Eighty-three bacteria, from a total of 1091, isolated from the rhizoplane of inoculated pasture clovers, inhibited the growth of two Rhizobium trifolii strains. The antagonists belonged to the genera Pseudomonas, Xanthomonas, Achromobacter, Flavobacterium, Alcaligenes, Erwinia, Aerobacter, Bacillus, Streptomyces, Nocardia, Corynebacterium, Arthrobacter, and Brevibacterium. Most, as well as the strongest, antagonists were confined to Pseudomonas. Two distinct groups were recognized within this genus. The A and B groups produced pink water soluble and green fluorescent pigments, respectively.

Activation of 6-phosphogluconate dehydrase by pyoverdine

Pseudomonas mildenbergii , when grown without iron, produces a green fluorescent pigment, pyoverdine ( Love & Hulcher, 1964), which has a molecular weight of 2,380 and consists of a peptide of threonine, serine, glutamic acid and lysine (4:2:1:1, molar ratio) with an N-methyl phenylacetyl hydroxamic acid group bound to the peptide (Hulcher, 1968). The present work defines a possible physiological role for pyoverdine as an activator of 6-phosphogluconate dehydrase.

Characteristics and Pathogenicity of a Capsulated Pseudomonas Isolated from Goldfish

Characteristics of a capsulated bacterium isolated from an epizootic among goldfish (Carassius auratus) were determined as well as the ability of the bacterium to produce experimental infections. The bacterium was found to be a gram-negative rod which oxidized carbohydrates, produced green fluorescent pigment, and otherwise seemed to fit into the genus Pseudomonas, except that it was nonmotile and failed to oxidize gluconate. These last two characteristics are typical of pseudomonads. However, the bacterium was classified as a pseudomonad, best fitting the description of the nonmotile variety of Pseudomonas fluorescens. Goldfish and rainbow trout (Salmo gairdneri) were infected experimentally by injection, but not by being fed bacteria-laden food. Goldfish were infected by exposure to the bacterium, but only if two or three scales were removed prior to exposure. It is suggested that the bacterium is an opportunistic pathogen.

CHARACTERISTICS AND PATHOGENICITY OF A CAPSULATED PSEUDOMONAS ISOLATED FROM GOLDFISH.

Characteristics of a capsulated bacterium isolated from an epizootic among goldfish (Carassius auratus) were determined as well as the ability of the bacterium to produce experimental infections. The bacterium was found to be a gram-negative rod which oxidized carbohydrates, produced green fluorescent pigment, and otherwise seemed to fit into the genus Pseudomonas, except that it was nonmotile and failed to oxidize gluconate. These last two characteristics are typical of pseudomonads. However, the bacterium was classified as a pseudomonad, best fitting the description of the nonmotile variety of Pseudomonas fluorescens. Goldfish and rainbow trout (Salmo gairdneri) were infected experimentally by injection, but not by being fed bacteria-laden food. Goldfish were infected by exposure to the bacterium, but only if two or three scales were removed prior to exposure. It is suggested that the bacterium is an opportunistic pathogen.

GREEN FLUORESCENT PIGMENT ACCUMULATED BY A MUTANT OF CELLVIBRIO GILVUS.

Love, Samuel H. (Bowman Gray School of Medicine, Winston-Salem, N.C.), and Frank H. Hulcher. Green fluorescent pigment accumulated by a mutant of Cellvibrio gilvus. J. Bacteriol. 87:39-45. 1964.-A mutant of Cellvibrio gilvus, designated strain 139A, liberated a green, fluorescent pigment into the surrounding culture medium. A study of the factors which affected the accumulation of this pigment led to the development of a chemically defined medium which supported maximal pigment accumulation in aerated, liquid cultures. d-Glucose, glycine or l-serine, l-phenylalanine, l-proline, and l-lysine comprised the organic components of this medium. The visible absorption spectrum of the pigment showed a maximal band at 400 mmu (pH 7.0). A difference spectrum between reduced and oxidized pigment showed loss of the band at 400 mmu upon oxidation. However, a methanol-extractable, flavinelike compound occurred in the wild strain but not in the mutant. Ferric ions added to the defined medium stimulated growth, with a concomitant reduction of pigment accumulation. Pigment was formed at a maximal rate during the stationary growth phase, and the highest yield was obtained by 18 hr. Organic solvents did not extract the pigment from water solutions. One and sometimes two, compounds absorbing at 400 mmu could be eluted by ion-exchange chromatography on Cellex-P (H(+)), which was used to separate the pigment from other components in the culture supernatants so that the radioactivity of the pigment could be measured. The mutant synthesized C(14)-labeled pigment from d-glucose-U-C(14) and from each of four amino acids (glycine-1-C(14), l-phenylalanine-U-C(14), l-proline-U-C(14), and l-lysine-U-C(14). Delta-Amino-levulenic acid-4-C(14) did not contribute C(14) to the pigment.