Effect of amino acids containing sulfur on dithiolopyrrolone antibiotic productions by Saccharothrix algeriensis NRRL B-24137

AIMS: To study the effect of sulfur-containing amino acids (L-cysteine, L-cystine, L-methionine and DL-ethionine) on the production of dithiolopyrrolone antibiotics by Saccharothrix algeriensis NRRL B-24137. METHODS AND RESULTS: The production levels of dithiolopyrrolones were investigated by using high performance liquid chromatography in a chemically semi-synthetic medium. The production of the studied antibiotics depends upon the nature, concentration and the time of addition of these sources in the culture medium. Both cysteine and cystine favoured the specific productions of dithiolopyrrolones; iso-butyryl-pyrrothine (ISP) by cysteine, however butanoyl-pyrrothine, senecioyl-pyrrothine and tigloyl-pyrrothine by cystine, when added initially to the culture medium. The maximum specific productions of dithiolopyrrolones were observed in the presence of 5 mmol l(-1) cystine for thiolutin, 5 mmol l(-1) cysteine for ISP, and 10 mmol l(-1) cystine for others studied dithiolopyrrolones as shown in Fig. 3. The production of these antibiotics was decreased when the concentrations of cysteine and cystine were in excess. All dithiolopyrrolone specific productions were strongly inhibited by addition of methionine and ethionine, without inhibition of mycelial growth. CONCLUSIONS: Among all studied amino acids, cystine and cysteine can be used as supplements for improvement the production of dithiolopyrrolone antibiotics by S. algeriensis NRRL B-24137. SIGNIFICANCE AND IMPACT OF THE STUDY: Dithiolopyrrolone antibiotics have many important applications for employing them as medicaments, particularly in the treatment of human and animal cancers. In the present work, the influence of containing-sulfur amino acids on dithiolopyrrolone antibiotic productions was studied. The obtained results can be employed for the optimization of the culture medium for the dithiolopyrrolone productions in higher quantities.     

Nutritional requirements for the production of dithiolopyrrolone antibiotics by Saccharothrix algeriensis NRRL B-24137

The amino acid and humic acid requirements of Saccharothrix algeriensis NRRL B-24137 for growth and production of the dithiolopyrrolone antibiotics were studied in a semi-synthetic medium (SSM). Nature and concentration of amino acids and humic acid strongly influenced the growth and dithiolopyrrolone specific production.

The highest value of thiolutin (acetyl-pyrrothine) specific production was obtained in the presence of 1 g/l humic acid (336 mg/g DCW), and in the presence of 5 mM l-cystine (309 mg/g DCW) as compared to 19 mg/g DCW obtained with the control. Furthermore, thiolutin production was increased about six-fold, four-fold and three-fold in the presence of l-proline, l-glutamic acid and dl-histidine, respectively. In contrast, the production of thiolutin was reduced by addition of other amino acids such as l-glutamine, dl-ethionine, l-methionine and l-arginine. The highest value of isobutyryl-pyrrothine production was obtained in the presence of 2,6-diaminopimelic acid and l-lysine (7.8 and 1.0 mg/g DCW, respectively). However, the highest value of butanoyl-pyrrothine production was obtained in the presence of humic acid (6.6 mg/g DCW), followed by l-cysteine and l-proline (3.6 and 3.2 mg/g DCW, respectively). In addition, the maximum specific production of senecioyl-pyrrothine (29 mg/g DCW) and tigloyl-pyrrothine (21 mg/g DCW) was obtained in the presence of humic acid. We found that, except for isobutyryl-pyrrothine, production of all dithiolopyrrolones was favoured by addition of l-proline. The maximum specific production was obtained with l-proline at concentrations of 2.50 mM for thiolutin (133 mg/g DCW), 1.25 mM for senecioyl-pyrrothine, tigloyl-pyrrothine and butanoyl-pyrrothine production (29, 23 and 3.9 mg/g DCW, respectively). Production of all dithiolopyrrolones strongly decreased as the l-methionine or dl-ethionine concentration was increased in the culture medium.     

The final acylation step in aromatic dithiolopyrrolone biosyntheses: Identification and characterization of the first bacterium N-benzoyltransferase from Saccharothrix algeriensis NRRL B-24137

The last step in the biosynthesis of dithiolopyrrolone antibiotics was thought to involve the transfer of acyl group from acyl-CoA to pyrrothine/holothin core. In Saccharothrix algeriensis NRRL B-24137, two acyltransferases, an acetyltransferase and a benzoyltransferase were proposed to catalyze this step. We have previously identified, in Sa. algeriensis genome, two open read frames, actA and actB patiently encoded these enzymes. This study focuses primarily on the characterization of the protein encoded by actA. After cloning and expressing of actA in Escherichia coli BL21, the recombinant protein encoded by actA was purified. Selectivity of ActA for pyrrothine/holothin as substrate and different acyl-CoA as co-substrate was evaluated using two acyls-groups, linear and aromatic. The enzyme was shown to prefer aromatic groups over linear groups as donor group; further neither product nor transfer was observed for linear groups. Therefore ActA has been determined to be a pyrrothine/holothin N-benzoyltransferase which can either pyrrothine (K_m of 72 μM) or holothin (K_m of 129.5 μM) as substrates and benzoyl-CoA (K_m of 348.65 and 395.28 μM) as co-substrates for pyrrothine and holothin, respectively. The optimum pH and temperature has been shown to be 8, 40 °C, respectively. ActA is the first enzyme characterized as N-benzoyltransferase in bacteria.     

New dithiolopyrrolone antibiotics induced by adding sorbic acid to the culture medium of Saccharothrix algeriensis NRRL B-24137

Dithiolopyrrolone antibiotics, produced by several microorganisms, are known for their strong antimicrobial activities. This class of antibiotics generated new interest after the discovery of their anticancer and antitumor properties. In this study, four new antibiotics were purified from the fermentation broth of Saccharothrix algeriensis NRRL B-24137 and characterized as dithiolopyrrolone derivatives. These new dithiolopyrrolone antibiotics were induced by adding sorbic acid, as precursor, at a concentration of 5 mM to the semi-synthetic medium. The analysis of the induced antibiotics was carried out by HPLC. The maximal production of the antibiotics PR2, PR8, PR9 and PR10 was 0.08±0.04, 0.21±0.04, 0.13±0.03 and 0.09±0.00 mg L(-1) , respectively, obtained after 8 days of fermentation. The chemical structures of these antibiotics were determined by (1) H- and (13) C-nuclear magnetic resonance, mass and UV-visible data. The four new dithiolopyrrolone antibiotics - PR2, PR8, PR9 and PR10 - were characterized, respectively, as crotonyl-pyrrothine, sorbyl-pyrrothine, 2-hexonyl-pyrrothine and 2-methyl-3-pentenyl-pyrrothine. The minimum inhibitory concentrations of the new induced antibiotics were determined.     

A stoichiometric reaction scheme for Saccharothrix algeriensis growth and thiolutin production

A new bacterial species, Saccharothrix algeriensis NRRL B-24137, was isolated in 1992 in the Sahara desert. This filamentous bacterium is able to produce dithiolopyrrolones, molecules presenting antibacterial, antifungal, and anticancer properties. In this study, a “reaction engineering” approach was adopted to gain more knowledge on the growth of Sa. algeriensis and its dithiolopyrrolone production on a semi-synthetic liquid medium. The objective is to establish a reaction scheme of the bacterium metabolism from extracellular experimental information, relatively easy to obtain. The approach enabled us to show that Sa. algeriensis could grow using several substrates that were sequentially consumed and that substrate limitation may induce a secondary metabolism in antibiotic production. From these qualitative data, a general reaction scheme was extracted consisting of four reactions: growth via amino acids, glucose consumption for maintenance, growth using glucose, and thiolutin production. The stoichiometric coefficients and the reaction extends were identified using a factorial analysis based on the bilinear structure of the component mass balances in a batch reactor. The analysis of the reaction stoichiometry enabled us to draw some conclusions concerning the substrate consumption pathway.     

Nutritional requirements for the production of dithiolopyrrolone antibiotics by Saccharothrix algeriensis NRRL B-24137

The amino acid and humic acid requirements of Saccharothrix algeriensis NRRL B-24137 for growth and production of the dithiolopyrrolone antibiotics were studied in a semi-synthetic medium (SSM). Nature and concentration of amino acids and humic acid strongly influenced the growth and dithiolopyrrolone specific production.The highest value of thiolutin (acetyl-pyrrothine) specific production was obtained in the presence of 1 g/l humic acid (336 mg/g DCW), and in the presence of 5 mM l-cystine (309 mg/g DCW) as compared to 19 mg/g DCW obtained with the control. Furthermore, thiolutin production was increased about six-fold, four-fold and three-fold in the presence of l-proline, l-glutamic acid and dl-histidine, respectively. In contrast, the production of thiolutin was reduced by addition of other amino acids such as l-glutamine, dl-ethionine, l-methionine and l-arginine. The highest value of isobutyryl-pyrrothine production was obtained in the presence of 2,6-diaminopimelic acid and l-lysine (7.8 and 1.0 mg/g DCW, respectively). However, the highest value of butanoyl-pyrrothine production was obtained in the presence of humic acid (6.6 mg/g DCW), followed by l-cysteine and l-proline (3.6 and 3.2 mg/g DCW, respectively). In addition, the maximum specific production of senecioyl-pyrrothine (29 mg/g DCW) and tigloyl-pyrrothine (21 mg/g DCW) was obtained in the presence of humic acid. We found that, except for isobutyryl-pyrrothine, production of all dithiolopyrrolones was favoured by addition of l-proline. The maximum specific production was obtained with l-proline at concentrations of 2.50 mM for thiolutin (133 mg/g DCW), 1.25 mM for senecioyl-pyrrothine, tigloyl-pyrrothine and butanoyl-pyrrothine production (29, 23 and 3.9 mg/g DCW, respectively). Production of all dithiolopyrrolones strongly decreased as the l-methionine or dl-ethionine concentration was increased in the culture medium.     

二硫吡咯酮类抗生素的生物合成与代谢工程应用北大核心CSCD

二硫吡咯酮类抗生素是一类具有独特的吡咯酮二硫杂环戊二烯(4H-[1,2]二硫[4,3-b]吡咯-5-酮)骨架的化合物的总称。基于N-7位酰基侧链的不同以及N-4位是否含有甲基,可分为N-methyl-Nacylpyrrothine、N-acylpyrrothine和thiomarinols等类别。迄今为止,已有27种该类化合物被报道,重要代表包括全霉素(holomycin)、硫藤黄菌素(thiolutin)、金霉素(aureothricin)以及最近发现的thiomarinols。就生物活性而言,二硫吡咯酮类抗生素具有广谱的抗细菌活性,对多种微生物,包括革兰氏阴性菌、革兰氏阳性菌以及寄生虫都有较好的杀灭活性。甚至一些二硫吡咯酮衍生物表现出较强的抗肿瘤活性。近几年来,多个二硫吡咯酮类抗生素的生物合成基因簇相继被报道,其生物合成机理也逐步被阐明。本文将针对目前国内外二硫吡咯酮类抗生素的生物合成研究进展,以及在组合生物合成与代谢工程领域所取得的成果进行综述,旨在为通过合成生物学的方法创造结构新颖、高效低毒的"非天然"二硫吡咯酮类化合物提供理论借鉴。     

Production of transglutaminase by Streptomyces isolates in solid-state fermentation

Aims:  To screen Streptomyces isolates for transglutaminase (TGase) production in solid-state fermentation (SSF) on various substrates.
Methods and Results:  Streptomyces mobaraensis NRRL B-3729, Streptomyces paucisporogenes ATCC 12596 and Streptomyces platensis NRRL 2364 strains were screened for extracellular TGase production in SSF on different substrates. High-protein-content beans, peas and lentils proved to be the best substrates. Good TGase production was obtained on liver kidney beans and green mung beans in a 4- to 6-day SSF. Temperature optima of the enzymes varied between 45 to 50°C. Molecular weight determined by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS PAGE) indicated similar size (∼37 kDa) for all three enzymes. TGase was the dominating protein band on SDS PAGE for two Streptomyces strains in SSF extracts. Other enzymes were present in smaller quantities.
Conclusions:  Streptomyces mobaraensis NRRL B-3729, S. paucisporogenes ATCC 12596 and S. platensis NRRL 2364 strains were successfully propagated under SSF conditions on crushed/milled liver kidney bean and green mung bean to obtain good level of TGase.
Significance and Impact of the Study:  Owing to much reduced production cost and direct applicability, SSF TGase without downstream processing (cheap in situ enzyme, crude enzyme) may be an excellent candidate for some nonfood applications.     

The Saharan isolate Saccharothrix algeriensis NRRL B-24137 induces systemic resistance in Arabidopsis thaliana seedlings against Botrytis cinerea

Background and aim

Saccharothrix algeriensis NRRL B-24137, isolated from a Saharan soil, has been described as a potential biocontrol agent against Botrytis cinerea and other phytopathogens. However, the plant protection mechanisms involved still need to be described. The aim of this study was to determine this protection phenomenon as well as parts of the mechanisms involved, using Arabidopsis thaliana seedlings and B. cinerea.

Methods

The bacterial colonization process was evaluated on A. thaliana seedlings using fluorescence in situ hybridization. Protection of A. thaliana seedlings inoculated with NRRL B-24137 against B. cinerea was then evaluated. Parts of the mechanisms involved in the systemic protection against B. cinerea were evaluated using known mutants of genes involved in jasmonate (JA)/ethylene (ET)/salicylic acid (SA) signaling. Other Arabidopsis mutants, AtrhbohD-3, AtrhbohF-3, and ups1-1 were also screened to determine other parts of the mechanisms involved.

Results

The results showed that the strain NRRL B-24137 colonized, epi- and endophytically, the roots of Arabidopsis seedlings but the strain was not a systemic colonizer during the time of the experiment. The strain NRRL B-24137 also reduced B. cinerea symptoms and the protection was linked to known mechanisms of induced systemic resistance (ISR; JA/ET signaling), as well as to functionality of AtrbohF oxidase and of UPS1. Crosstalk between ET/JA and SA signaling could also be involved.

Conclusions

The isolate NRRL B-24137, after colonizing the root systems of A. thaliana, induces an ISR against B. cinerea, which is JA/ET dependent, but could also require SA crosstalk and protection could also require NAPDH oxidases and UPS1 functionalities.     

Alternansucrase mutants of Leuconostoc mesenteroides strain NRRL B-21138

Alternan is a unique α-D-glucan of potential commercial interest, produced by rare strains of Leuconostoc mesenteroides. Natural isolates that produce alternan, such as NRRL B-1355, also produce dextran as a troublesome contaminant. We previously isolated mutants of strain NRRL B-1355 that are deficient in dextran production, including the highly stable strain NRRL B-21138. In the current work, we mutagenized strain NRRL B-21138 and screened survivors for further alterations in production of alternansucrase, the enzyme that catalyzes the synthesis of alternan from sucrose. Second generation mutants included highly stable strain NRRL B-21297, which produced four-fold elevated levels of alternansucrase without an increase in the proportion of dextransucrase activity. Such alternansucrase overproducing strains will facilitate studies of this enzyme, and may become valuable for the enzymatic production of alternan. Another highly stable mutant strain, NRRL B-21414, grew slowly on sucrose with negligible production of glucan or extracellular glucansucrase activity. This strain may prove useful as an expression host for glucansucrase genes. Received 30 July 1996/ Accepted in revised form 15 December 1996     

Yield enhancement strategy of dithiolopyrrolone from Saccharothrix algeriensis by aliphatic alcohols supplementation

The production of dithiolopyrrolones by Saccharothrix algeriensis was investigated after supplementing the culture medium with ethanol and/or 1-butanol. Optimal conditions for the addition of ethanol to the culture medium provided a maximal dithiolopyrrolone titer of about 200 mg⋅L⁻¹ after 5 days of culture, roughly corresponding to a 600%-increase. Using NAD(P)H oxidase inhibitor (diphenyleneiodonium) or reactive oxygen species scavenger (para-aminobenzoic acid), we suppose that ethanol promotes the formation of reactive oxygen species in Saccharothrix algeriensis, which, in turn, could induce biomass decline and dithiolopyrrolone overproduction. However, the underlying mechanisms remain to be elucidated. These results may be helpful for the control of dithiolopyrrolone yields from Saccharothrix algeriensis cultures.     

Leuconostoc mesenteroides glucansucrase synthesis of flavonoid glucosides by acceptor reactions in aqueous-organic solvents

The enzymatic glucosylation of luteolin was attempted using two glucansucrases: the dextransucrase from Leuconostoc mesenteroides NRRL B-512F and the alternansucrase from L. mesenteroides NRRL B-23192. Reactions were carried out in aqueous-organic solvents to improve luteolin solubility. A molar conversion of 44% was achieved after 24h of reaction catalysed by dextransucrase from L. mesenteroides NRRL B-512F in a mixture of acetate buffer (70%)/bis(2-methoxyethyl) ether (30%). Two products were characterised by nuclear magnetic resonance (NMR) spectroscopy: luteolin-3'-O-alpha-d-glucopyranoside and luteolin-4'-O-alpha-d-glucopyranoside. In the presence of alternansucrase from L. mesenteroides NRRL B-23192, three additional products were obtained with a luteolin conversion of 8%. Both enzymes were also able to glucosylate quercetin and myricetin with conversion of 4% and 49%, respectively.     

Influence on dithiolopyrrolone antibiotic production by organic acids in Saccharothrix algeriensis NRRL B-24137

The influence of organic acids on growth and dithiolopyrrolone antibiotic production by Saccharothrix algeriensis NRRL B-24137 was studied. The production of dithiolopyrrolones depends upon the nature and concentration of the organic acids in the culture medium. Study of the nature of organic acids showed that the most effective organic acids for thiolutin specific production were maleic, 4-hydroxybenzoic, benzentetracarboxylic, pantothenic, pivalic and pyruvic acids (which yielded almost five-fold over the starting medium) and pimelic acid (more than three-fold). 4-Bromobenzoic acid showed the best production of senecioyl-pyrrothine (59 mg g⁻¹ DCW). Tiglic acid showed the best production of tigloyl-pyrrothine (22 mg g⁻¹ DCW). The highest yield of isobutyryl-pyrrothine (7.6 mg g⁻¹ DCW) was observed in the presence of crotonic acid. Sorbic acid yielded the best production of butanoyl-pyrrothine (26 mg g⁻¹ DCW). Methacrylic, butyric, pyruvic and 4-bromobenzoic acids also exhibited the best production of butanoyl-pyrrothine (27–11-fold).Study of organic acid concentration showed that among the selected organic acids, pimelic acid yielded the highest specific production of thiolutin (91 mg g⁻¹ DCW) at 7.5 mM; and senecioyl-pyrrothine (11 mg g⁻¹ DCW), tigloyl-pyrrothine (9 mg g⁻¹ DCW) and butanoyl-pyrrothine (3.5 mg g⁻¹ DCW) at 5 mM. Pyruvic acid at 1.25 mM enhanced the production of senecioyl-pyrrothine (4.3 mg g⁻¹ DCW). The maximum production of tigloyl-pyrrothine (18.6 mg g⁻¹ DCW) was observed in the presence of tiglic acid at 2.5 mM. Maximum production of isobutyryl-pyrrothine was observed in the presence of 7.5 mM tiglic acid. In addition, methacrylic acid (at 5 mM) and butyric acid (at 2.5 mM) enhanced the production of butanoyl-pyrrothine (26 and 20 times, respectively).The above results can be employed in the optimisation of the culture medium for the production of dithiolopyrrolone in higher quantities.     

Biosynthesis of spectinomycin: heterologous production of spectinomycin and spectinamine in an aminoglycoside-deficient host, Streptomyces venezuelae YJ003

Aims:  To obtain spectinomycin and spectinamine by heterologous expression into the biosynthetic deoxysugar (desosamine) gene-deleted host Streptomyces venezuelae YJ003.
Methods and Results:  The 17-kb spectinomycin biosynthetic gene cluster from Streptomyces spectabilis ATCC 27741 was heterologously expressed into Streptomyces venezuelae YJ003. Furthermore, the speA , speB and spcS2 encoded in the spectinomycin biosynthetic gene cluster of cosmid pSPC8 were also heterologously characterized to be responsible for the production of spectinamine.
Conclusions:  The results of this study indicated that pSPC8 contains all the genes necessary for the biosynthesis of spectinomycin. We also concluded that SpeA, SpeB and SpcS2 are sufficient for the biosynthesis of spectinamine. We also verified that SpeB and SpcS2 show dual character in the biosynthetic pathway of spectinomycin in Streptomyces spectabilis .
Significance and Impact of the Study:  This is the report regarding the expression of a biosynthetic gene cluster that gives rise to the production of aminoglycoside antibiotics in Streptomyces venezuelae YJ003. Therefore, this work may serve as a foundation for further research on spectinomycin biosynthesis and other aminoglycosides.     

Electrochemical checking of aerobic isolates from electrochemically active biofilms formed in compost

Aims:  To design a cyclic voltammetry (CV) procedure to check the electrochemical activity of bacterial isolates that may explain the electrochemical properties of biofilms formed in compost.
Methods and Results:  Bacteria catalysing acetate oxidation in garden compost were able to form electrochemically active biofilms by transferring electrons to an electrode under chronoamperometry. They were recovered from the electrode surface and identification of the isolates using 16S rRNA sequencing showed that most of them were Gammaproteobacteria, mainly related to Enterobacter and Pseudomonas spp. A CV procedure was designed to check the electrochemical activity of both groups of isolates. Preliminary CVs suggested that the bacteria were not responsible for the catalysis of acetate oxidation. In contrast, both groups of isolates were found to catalyse the electrochemical reduction of oxygen under experimental conditions that favoured adsorption of the microbial cells on the electrode surface.
Conclusions:  Members of the genera Enterobacter and Pseudomonas were found to be able to catalyse the electrochemical reduction of oxygen.
Significance and Impact of the Study:  This study has shown the unexpected efficiency of Enterobacter and Pseudomonas spp. in catalysing the reduction of oxygen, suggesting a possible involvement of these species in biocorrosion, or possible application of these strains in designing bio-cathode for microbial fuel cells.     

Molecular cloning and functional characterization of a histidine decarboxylase from Staphylococcus capitis

Aims:  Histamine intoxication is probably the best known toxicological problem of food-borne disease. A histamine-producing Staphylococcus capitis strain has been isolated from a cured meat product. The aim of this study was to gain deeper insights into the genetic determinants for histamine production in Staph. capitis .
Methods and Results:  The nucleotide sequence of a 6446-bp chromosomal DNA fragment containing the hdcA gene encoding histidine decarboxylase (HDC) has been determined in Staph. capitis IFIJ12. This DNA fragment contains five complete and two partial open reading frames. Putative functions have been assigned to gene products by sequence comparison with proteins included in the databases. The hdcA gene has been expressed in Escherichia coli resulting in HDC activity. The presence of a functional promoter (P hdc ) located upstream of hdcA has been demonstrated. Insertion of the histamine biosynthetic locus in Staph. capitis seems to be associated with a noticeable genome reorganization.
Conclusions:  Among the staphylococcal species analysed in this study only Staph. capitis strains produce histamine. The hdcA gene cloned from Staph. capitis encodes a functional HDC that produce histamine from the amino acid histidine.
Significance and Impact of the Study:  The identification of the DNA region involved in histamine production in Staph. capitis will allow further work in order to avoid histamine production in foods.     

Conversion of Unsaturated Fatty Acids by Bacteria Isolated from Compost

A compost mixture amended with soybean oil was enriched in microorganisms that transformed unsaturated fatty acids (UFAs). When oleic acid or 10-ketostearic acid was the selective fatty acid, Sphingobacterium thalpophilum (NRRL B-23206, NRRL B-23208, NRRL B-23209, NRRL B-23210, NRRL B-23211, NRRL B-23212), Acinetobacter spp. (NRRL B-23207, NRRL B-23213), and Enterobacter cloacae (NRRL B-23264, NRRL B-23265, NRRL B-23266) represented isolates that produced either hydroxystearic acid, ketostearic acid, or incomplete decarboxylations. When ricinoleic (12-hydroxy-9-octadecenoic) acid was the selective UFA, Enterobacter cloacae (NRRL B-23257, NRRL B-23267) and Escherichia sp. (NRRL B-23259) produced 12-C and 14-C homologous compounds, and Pseudomonas aeruginosa (NRRL B-23256, NRRL B-23260) converted ricinoleate to a trihydroxyoctadecenoate product. Also, various Enterobacter, Pseudomonas, and Serratia spp. appeared to decarboxylate linoleate substrate incompletely. These saprophytic, compost bacteria were aerobic or facultative anaerobic Gram-negative and decomposed UFAs through decarboxylation, hydroxylation, and hydroperoxidation mechanisms. Received: 3 November 1998 / Accepted: 30 November 1998     

Diversity of Oleic Acid,Ricinoleic Acid and Linoleic Acid Conversions Among <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> Strains*

Sixteen Pseudomonas aeruginosa strains, including patent strain NRRL B-18602, three recent isolates from composted materials amended with ricinoleic acid, and 12 randomly selected from the holdings of the ARS Culture Collection, were examined for their fatty acid converting abilities. The study examined the bioconversion of oleic acid to 7,10-dihydroxy-8(E)-octadecenoic acid (DOD) and ricinoleic acid to 7,10,12-trihydroxy-8(E)-octadecenoic acid (TOD). A new DOD-like compound from linoleic acid was observed. All strains except NRRL B-247 exhibited varying levels of DOD production. NRRL B-1000, NRRL B-18602 and NRRL B-23258 with yields up to 84% were among the best DOD producers. TOD production generally paralleled DOD production at a relatively lower yield of up to 15%. Strains NRRL B-1000 and NRRL B-23260 were the best TOD producers. A DOD-like product in low yields was obtained from linoleic acid. The fatty acid bioconversion capability was related neither to growth rate nor to variation in the greenish pigmentation of the strains. Production of significant quantities of DOD and TOD from oleic and ricinoleic acids, respectively, appeared to be a characteristic trait of P. aeruginosa strains. A number of highly effective strains for DOD production were identified.     

Phylogeny of Marine Bacillus Isolates from the Gulf of Mexico

The phylogeny of 11 pigmented, aerobic, spore-forming isolates from marine sources was studied. Forty-two biochemical characteristics were examined, and a 16S rDNA sequence was obtained for each isolate. In a phylogenetic tree based on 16S sequencing, four isolates (NRRL B-14850, NRRL B-14904, NRRL B-14907, and NRRL B-14908) clustered with B. subtilis and related organisms; NRRL B-14907 was closely related to B. amyloliquefaciens. NRRL B-14907 and NRRL B-14908 were phenotypically similar to B. amyloliquefaciens and B. pumilus, respectively. Three strains (NRRL B-14906, NRRL B-14910, and NRRL B-14911) clustered in a clade that included B. firmus, B. lentus, and B. megaterium. NRRL B-14910 was closely related phenotypically and phylogenetically to B. megaterium. NRRL B-14905 clustered with the mesophilic round spore-producing species, B. fusiformis and B. sphaericus; the isolate was more closely related to B. fusiformis. NRRL B-14905 displayed characteristics typical of the B. sphaericus-like organisms. NRRL B-14909 and NRRL B-14912 clustered with the Paenibacillus species and displayed characteristics typical of the genus. Only NRRL B-14851, an unusually thin rod that forms very small spores, may represent a new Bacillus species. Received: 8 December 1999 / Accepted: 14 February 2000