Response of growth and fatty acid compositions of Nannochloropsis sp. to environmental factors under elevated CO2 concentration

Nannochloropsis sp. was grown with different levels of nitrate, phosphate, salinity and temperature with CO₂ at 2,800 μl l⁻¹. Increased levels of NaNO₃ and KH₂PO₄ raised protein and polyunsaturated fatty acids (PUFAs) contents but decreased carbohydrate, total lipid and total fatty acids (TFA) contents. Nannochloropsis sp. grew well at salinities from 22 to 49 g l⁻¹, and lowering salinity enhanced TFA and PUFAs contents. TFA contents increased with the increasing temperature but PUFAs contents decreased. The highest eicosapentaenoic acid (EPA, 20:5ω3) content based on the dry mass was above 3% under low N (150 μM NaNO₃) or high N (3000 μM NaNO₃) condition. Excessive nitrate, low salinity and temperature are thus favorable factors for improving EPA yields in Nannochloropsis sp.     

Improved production of various polyunsaturated fatty acids through filamentous fungus Mortierella alpina breeding

Studies on the application of functional lipids such as polyunsaturated fatty acids (PUFAs) have proceeded in various fields regarding health and dietary requirements in a search for novel and rich sources. Filamentous fungus Mortierella alpina 1S-4 produces triacylglycerols rich in arachidonic acid, ones reaching 20 g/L and containing 30–70% arachidonic acid as to the total fatty acids. Mutants derived from M. alpina 1S-4, defective in Δ5 and Δ6 desaturases, accumulate triacylglycerols rich in unique PUFAs, i.e., dihomo-γ-linolenic acid and Mead acid, respectively. Furthermore, various mutants derived from M. alpina 1S-4 have led to the production of oils containing n−1, n−3, n−4, n−6, n−7, and n−9 PUFAs. A variety of genes encoding fatty acid desaturases and elongases involved in PUFA biosynthesis in M. alpina 1S-4 has been isolated and characterized. Molecular breeding of M. alpina strains by means of manipulation of these genes facilitates improvement of PUFA productivity and elucidation of the functions of enzymes involved in PUFA biosynthesis.     

乳酸菌的基因组编辑技术研究进展

乳球菌(Lactococcussp.)和乳杆菌(Lactobacillussp.)是工业上常用的乳酸菌(lacticacid bacteria,LAB),长期应用于食品和饮料的发酵。近年来,随着分子操作及遗传改造技术的不断完善,推动了乳球菌和乳杆菌的基础和应用研究,其作为功能菌株和工业微生物细胞工厂的重要潜能也不断突显出来。本文综述了工业常用乳酸菌的基因组编辑技术研究进展,着重介绍了基于整合质粒的敲除、敲入,基于基因组重组工程的精细修饰和敲除、敲入,以及基于成簇的规律性间隔的短回文重复序列及其相关蛋白9[(clusteredregularlyinterspacedshortpalindromicrepeats(CRISPR)/CRISPR-associated nuclease9 (Cas9), CRISPR/Cas9]系统的基因组编辑技术。     

Variability of hydrocarbon and fatty acid components in cultures of the filamentous cyanobacterium Scytonema sp. isolated from microbial community "black cover" of limestone walls in Jerusalem

The hydrocarbon and lipid components of four strains of the filamentous cyanobacterium Scytonema sp. isolated from microbial community Black Cover of limestone walls in Jerusalem were identified by gas chromatography–mass spectrometry using serially coupled capillary columns. The dominant compounds were: 1-heptadecyne (1.5-8%), hexadecanoic acid (14-36%), (Z,Z)-9,12-octadecadienoic acid (12-30%), (Z,Z,Z)-9,12,15-octadecatrienoic acid (6-12%), n-heptadecane (4-16%), and 1-heptadecene (1.5-8%). In addition to unsaturated alkanes and fatty acids, the very long-chain (C₃₀-C₃₂) hydrocarbons, squalene (2.4-3.0%), and branched 4,8,12-trimethyl-C_13:0 acid were also isolated. Two major hydrocarbons were detected in the cyanobacteria species using GC-MS and ¹³C-NMR.     

Detection of high-phosphorus tolerant VAM-fungi colonizing hops and peppermint

Summary In a continuing search for field sources of endomycorrhizal fungi that tolerate high levels of available phosphorus in soil, samples were obtained from long-established liberally fertilized hops (Humulus lupulus L.) and peppermint (Mentha piperita L.) fields. In hops fields, where extractable P (Bray) levels ranged from 21–196 ppm, spores of twoAcaulospora spp. and sixGlomus spp. were isolated. From peppermint fields with P levels of 44 to 244 ppm, spores of oneAcaulospora sp., fourGlomus spp. and oneGigaspora sp. were obtained. Vesicular-arbuscular mycorrhizal (VAM) fungi were found colonizing most roots to some extent, despite the very high fertility of several sites. Although percentages of colonization in hops tended to be low, ranging from 0 to 9.3%, colonizations in peppermint roots were considerably higher averaging 26.5% between the sampled fields, notwithstanding P levels as high as 244 ppm. Curiously, considerable external VAM hyphae were found adherent to or entangling roots in many cases, even where percentage of root length with internal VAM structures was low. The functional benefit, neutrality, or detriment to plants of such high-P tolerant species remains to be determined.Contribution from the USDA-ARS, Horticultural Crops Research Laboratory and the Dept. of Soil Science, Oregon State University, in cooperation with the Oregon State University Agricultural Experiment Station, Corvallis, OR 97331. Technical paper no. 6979 of the latter     

Improvement of arachidonic acid and eicosapentaenoic acid production by increasing the copy number of the genes encoding fatty acid desaturase and elongase into <Emphasis Type="Italic">Pichia pastoris</Emphasis>

Genes encoding Δ6 desaturase, Δ6 fatty acid elongase, and Δ5 desaturase from the alga, Phaeodactylum tricornutum, were co-expressed in Pichia pastoris to produce arachidonic acid (ARA; 20:4 Δ^{5, 8, 11, 14}) and eicosapentaenoic acid (EPA; 20:5 Δ^{5, 8, 11, 14, 17}). A panel of Pichia clones carrying progressively increasing copies of the heterologous gene expression cassette was created using an in vitro multimerization approach. ARA and EPA accumulated up to 0.3 and 0.1% of total fatty acids, respectively, in the recombinant P. pastoris carrying with double copies of these three heterologous genes, as compared to 0.1 and 0.05%, respectively, in the recombinant P. pastoris with single copy. Yun-Tao Li and Mao-Teng Li contributed equally to this work.     

Fatty acid composition of selected prosthecate bacteria

The cellular fatty acid composition of 14 strains of Caulobacter species and types, two species of Prosthecomicrobium, and two species of Asticcacaulis was determined by gas-liquid chromatography. In most of these bacteria, the major fatty acids were octadecenoic acid (C18:1), hexadecenoic acid (C16:1) and hexadecanoic acid (C16:0). Some cyclopropane and branched chain fatty acids were detected in addition to the straight chained acids. Hydroxytetradecanoic acid was an important component of P. enhydrum but significant amounts of hydroxy acids were not detected in other prosthecate bacteria examined.Abbreviations DEGS diethylene glycol succinate - A measare of association Dedicated to Prof. R. Y. Stanier on the occasion of his 60th birthday     

Effect of growth temperature on the fatty acid composition of Mycobacterium phlei

In Mycobacterium phlei, fatty acid unsaturation increased with decreasing temperature. The 10-hexadecenoic acid content increased as the temperature was reduced from 35°C to 26–20°C. At lower temperatures tuberculostearic acid decreased while oleic and linoleic acids increased, the latter being found in M. phlei for the first time. Concomitantly palmitic acid content decreased, and the 6- and 9-hexadecenoic acids increased slightly on reducing the temperature from 35 to 10°C. Thus, down to 26–20°C palmitic acid was mainly replaced by 10-hexadecenoic acid. From this range down to 10°C, palmitic and tuberculostearic acids were replaced by oleic and linoleic acids. Consequently, fatty acid branching decreased and mean chain length increased, as the temperature was reduced. These observations support the view that regulation of membrane fatty acid composition is part of microbial temperature adaptation, and that themechanism behind the responses might be more complex than generally believed.Abbreviations ACP acyl carrier protein - FAS I (Type I) fatty acid synthetase I - FAS II (Type II) fatty acid synthetase II - MGLP methylglucose containing lipopolysaccharide - MMP methylmannose containning polysaccharide     

Identification and characterization of lactic acid bacteria in ragi tape

One hundred and eighteen lactic acid bacteria (LAB) were isolated from five different types of ragi tape, a traditional dry-starter of Balinese rice wine. The isolates could be classified into three groups based on the cell shape and capability to produce gas from glucose. Group I contained 66 homofermentative cocci, group II contained seven homofermentative rods, and group III contained 45 heterofermentative rods. Among these 118 isolates, 21 isolates representing these groups were selected and were first identified using phenotypic characters. The identification performed phenotypically was confirmed by sequencing of variable region 8 (V8) of the 16S rDNA. The comparative studies led to the identification of Pediococcus pentosaceus, Enterococcus faecium, Lactobacillus curvatus, Weissella confusa, and W. paramesenteroides from the ragi tape examined.     

Changes in fatty acid composition in Pseudomonas putida and Pseudomonas stutzeri during naphthalene degradation

The effects of naphthalene on the whole cell-derived fatty acid composition of Pseudomonas putida and Pseudomonas stutzeri during naphthalene degradation were investigated. These strains differed in their abilities to degrade naphthalene and in 1,2-catechol dioxygenase activities. The cells of both strains reacted to the addition of naphthalene with an increase in the saturated/unsaturated ratio. The dynamic changes comprised also alterations in the percentage of hydroxy, cyclopropane and branched fatty acids. Upon the exposure of naphthalene, new fatty acids were detected.     

Fatty acid composition of various hyphal budding bacteria

The total fatty acid composition of various strains of Hyphomicrobium, Pedomicrobium, and Rhodomicrobium spp. was determined by gas chromatography. In addition, the fatty acid pattern of a new hyphal budding bacterium, strain F-1, was compared with the other patterns obtained. Octadecenoic acid was the main component in most strains, comprising up to 75% of the total fatty acids. Lactobacillic acid and 3-methoxy-tetradecanoic acid were present in varying amounts in the lipids of all organisms except for the new isolate, F-1. This latter strain contained, however, large amounts of iso-heptadecanoic and iso-heptadecenoic acids, not present in the other budding bacteria studied. This composition was consistently found under various culture conditions. The data indicate that, except for the new bacterium F-1, the hyphal budding bacteria studied here are closely related. The total fatty acid composition is thought to be a useful taxonomic criterion for differentiation of these bacteria.     

Elimination of n-butylated hydroxytoluene methylation during fatty acid analysis by gas chromatography

An improved method for fatty acids analysis with optimum recovery of highly polyunsaturated fatty acids methyl esters in biological systems is presented. The method is based on transesterification of phospholipid and triacylglycerols to fatty acid methyl esters using a commercially available reagent, Methyl-Prep II. Without proper precautions, as much as 50% of n-butylated hydroxytoluene (BHT) added to prevent oxidation of polyunsaturated fatty acids, could be methylated during the transesterification step. Methylated BHT elutes close to 14:0 (myristic acid) and no longer functions as an antioxidant, but the modified conditions virtually eliminate the methylation of BHT. Sample extraction and methylation was completed in 30 min at room temperature. A chelator (diethylenetriamine-pentaacetic acid; DTPA) is also added to prevent peroxidation of metal catalyzed free radical chain reactions. The standard deviations of the major fatty acids from multiple human plasma samples prepared on different days were less than 5%. The recovery of arachidonic acid, 20:4, from plasma was improved using the new method, and the recovery for docosahexaenoic acid, 22:6, spiked to human plasma was found to be 99%.     

Unsaturated fatty acid composition and biosynthesis in Oscillatoria limnetica and other cyanobacteria

A number of cyanobacteria showing a high degree of adaptation to life under reduced oxygen tensions as witnessed by their potency of facultative anoxygenic CO₂ photoassimilation with sulfide as electron donor were found to lack polyunsaturated fatty acids in their lipids. Lack of polyunsaturated fatty acids was found in representatives of different taxonomic groups. One of the strains lacking polyenoic acids was Oscillatoria limnetica, which can alternatively grow acrobically or anaerobically with sulfide as electron donor. This organism was found to synthesize monounsaturated fatty acids by desaturation of their saturated counterparts, in the presence as well as in the absence of molecular oxygen.Abbreviations ACP Acyl carrier protein - DCMU 3(3,4-dichlorophenyl)-1,1-dimethylurea     

Effect of growth phase on the fatty acid composition of Thermus spp.

Two yellow and two red pigmented strains of Thermus were monitored for changes in fatty acid content and composition with reference to growth phase at the optimum temperature. Fatty acid content per mg of dry weight increased as the cultures aged. In addition the quantities of iso C 15:0, iso C 17:0 and iso C 16:0 increased in yellow pigmented strains, but in red pigmented strains, an increase was seen in iso C 15:0, but C 16:0 and iso C 16:0 levels decreased. Thus the fatty acid composition of these organisms varies with growth phase, and shows also strain specific variability.     

Alterations in growth temperature range and fatty acid composition of Thermus as a result of plasmid elimination

Elimination of plasmids from Thermus flavus, T. thermophilus and three wild Thermus strains caused alterations in growth temperature range, pigmentation and membrane fatty acids without affecting viability. Following plasmid elimination all Thermus strains lost their ability to grow above 70°C. In addition, the minimum growth temperature was lowered by 5–10°C. Fatty acids were reduced by an average of approximately 35%. In addition, the contribution of iso- and anteisobranched fatty acids were altered in four of the five strains. The iso C_15:0/iso C_17:0 ratio approached 1.0 in all strains, whereas the anteiso C_15:0/anteiso C_17:0 was reduced to 0.2. The iso C_16:0/normal-C_16:0 ratio increased in all strains due to an increase in iso C_16:0 in four strains and a reduction in normal-C_16:0 relative to iso C_16:0 in one strain. However, it was evident that the plasmid-free strains were able to compensate for these alterations in membrane fluidity to a certain extent by reducing the average chain length of isobranched acids. Altered fatty acid metabolism at the level of precursors may have influenced membrane composition and consequently growth temperature range.     

Synthesis in vitro of very long chain fatty acids in Vibrio sp. strain ABE-1

The activity of fatty acid synthetase (FAS) from Vibrio sp. strain ABE-1 required the presence of acyl carrier protein and was completely inhibited by thiolactomycin, an inhibitor specific for a type II FAS. These observations indicate that this enzyme is a type II FAS. Analysis by gas-liquid chromotography of the reaction products synthesized in vitro from [2-¹⁴C]malonyl-CoA by the partially purified FAS revealed, in addition to 16-and 18-carbon fatty acids which are normal constituents of this bacterium, the presence of fatty acids with very long chains. These fatty acids were identified as saturated and mono-unsaturated fatty acids with 20 up to as many as 30 carbon atoms. The longest fatty acids normally found in this bacterium contain 18-carbon atoms. These results suggest that the FAS from Vibrio sp. strain ABE-1 has potentially the ability to synthesize fatty acids with very long chains.Abbreviations ACP acyl carrier protein - FAME fatty acid methyl ester - FAS fatty acid synthetase - FID flame ionization detection - GLC gas-liquid chromatography - TLC thin-layer chromatography - In designations of fatty acids, such as 16:0, 16:1, etc the colon separates the number that denotes the number of carbon atoms and the number that denotes the number of double bonds, respectively, in the molecule - 16:0-CoA CoA ester of 16:0     

Polar lipids and fatty acid composition ofThermus strains from New Zealand

The polar lipids and fatty acid composition ofThermus aquaticus YT-1 and YS 041,T. filiformis Wai33 A1 and eighteen isolates from New Zealand, several of which are attributed toT. filiformis, were compared to complement the taxonomy of these organisms. The polar lipid patterns were essentially similar in all strains and consisted of one major phospholipid and one major glycolipid. The fatty acid analysis produced three basic groups corresponding toT. filiformis Wai33 A1,T. aquaticus and the third to the other New Zealand strains. The presence of hydroxy fatty acids is reported inThermus spp. for the first time.     

Characterization of disomic addition lines Brassica napus-Brassica nigra by isozyme,fatty acid,and RFLP markers

Summary Six Brassica napus — B. nigra disomic addition lines were characterized by isozyme, fatty acid, and RFLP markers. The markers were arranged in six synteny groups, representing six of the eight chromosomes present in the B. nigra genome. Synteny group 1 displayed high levels of linoleic and linolenic acids in the seeds of the B. nigra parent. Synteny group 3 accumulated higher levels of eicosenoic and erucic acid than B. nigra. Three of the lines transmitted the alien chromosome to 100% of the progeny. The rest had variable transmission rates but all were above 50%. Most of the lines produced disomic addition plants in their progeny, suggesting pollen transmission of the alien chromosome. In addition to the marked lines, six others remained unmarked. These could be grouped into two classes according to their alien chromosome transmission. It is likely that they represent the two other B. nigra chromosomes that remained uncharacterized by the markers. No diploid individuals carrying B. nigra genome-specific markers were detected in the progenies studied.     

Settlement and post-settlement processes limit the abundance of the geniculate coralline alga Calliarthron on subtidal walls

Cover of geniculate coralline algae is significantly greater on subtidal platforms than on vertical walls in a central California kelp forest. To determine the cause(s) of this pattern, settlement and growth of Calliarthron were compared on horizontal and vertical substrata attached to platforms or walls. Recruitment on horizontal substrata attached to reef walls indicated that spores were abundant along the walls. Densities of recruits were significantly smaller on vertical substrata attached to walls than on horizontal substrata attached to platforms, indicating that Calliarthron settlement is reduced on vertical substrata. Growth of basal crusts and the densities of fronds were also significantly greater on horizontal substrata attached to platforms than on vertical substrata attached to walls. Irradiance reaching wall surfaces was less than irradiance reaching platform surfaces. We suggest that light reaching wall surface is low enough to affect Calliarthron growth. These results indicate that the low abundance of Calliarthron on subtidal walls is a result of poor recruitment and slow growth.