Effect of pH and age of culture on cellular fatty acid composition of Leuconostoc oenos

Z. DRICI-CACHON, J.F. CAVIN AND C. DIVIÈS. 1996. This study is concerned with the fatty acid composition of three strains of Leuconostoc oenos grown at different p^H. The most abundant fatty acids were C18: 1 w ⁹, C19: 0 cy( w ^9,10) and C16:0, followed by C16: 1 w ⁹ and C14: 0. The p^H considerably modified the fatty acid distribution in Lo107 (an acidophilic strain) and Lo8413 (a moderately acidophilic strain). However, moderate changes occurred for LoATCC 23277 (a less acidophilic strain). At p^H 2.9, Lo107 has a remarkably high level of C19: 0 cy-( w ^9,10) and C19:0 cy( w ^11,12). Proportions of C18: 1 and C19:0 cyclo acids varied mainly with the p^H of the medium and also as a function of growth phase. The degree of unsaturation of fatty acids also varied with p^H.     

The Effect of Temperature on the Fatty Acid Composition of Tetrahymena pyriformis WH-14

SYNOPSIS. A reduction in the growth temperature of Tetrahymena pyriformis strain WH-14 from 35 C to 15 C resulted in distinct alterations in the fatty acid composition of the glycerophospholipids. The proportion of normal saturated acids declined from 26 to 19%; palmitoleic acid increased by 6%, and the composition of the polyunsaturated fatty acids increased in 18:2 Δ^6,11(n) and decreased in 18:2 Δ^9,12(n) and 18:3 Δ^6,9,12(n). The unsaturation index (the average number of double bonds/100 molecules) did not change with a shift in temperature.
Two biosynthetic pathways exist in Tetrahymena for the formation of unsaturated fatty acids. The observed changes in fatty acid composition that accompany a lowering of the environmental temperature can be accounted for by a reduction in the accumulation of products of the fatty acid pathway leading to the formation of γ-linolenic acid [16:0(n) → 18:0(n) → 18:1 Δ⁹(n) → 18:2 Δ^9,12(n) → 18:3 Δ^6,9,12(n)] and an increase in the components of the pathway leading to the formation of 18:2 Δ^6,11(n) [16:0(n) → 16:1 Δ⁹(n) → 18:1 Δ¹¹(n) → 18:2 Δ^6,11(n)]. The data suggest that the regulatory mechanism in Tetrahymena differs from that found in some bacteria where a simple substitution of unsaturated fatty acids for saturated fatty acids occurs at low culture temperatures.     

Evidence for the synthesis of the multi-positional isomers of monounsaturated fatty acid in Methylococcus capsusatus by the anaerobic pathway

Abstract The biosynthesis of the positional isomers of the monounsaturated fatty acids of Methylococcus capsulatus (Bath) has been investigated by studying the incorporation of [2-¹⁴C]malonyl CoA into long-chain fatty acids in vitro. The major unsaturated products were Δ ⁹ 16:1 and Δ ¹¹ 18:1; however, Δ ⁸, Δ ¹⁰ and Δ ¹¹ 16:1, as well as, Δ ¹⁰, Δ ¹² and Δ ¹³ 18:1 were also synthesized. The exclusion of O₂ from the reaction vessel did not affect the synthesis of unsaturated fatty acids or the double bonds positions. Cerulenin inhibited the synthesis of unsaturated fatty acid more than saturated fatty acid. The use of both [1-¹⁴C] octanoate and [1-¹⁴C] decanoate as substrate resulted in the synthesis of long-chain fatty acids, however, unsaturates were only synthesized from octanoate. These results imply that the unique positional isomers of M. capsulatus are not synthesized by an aerobic mechanism.     

Nisin induces changes in membrane fatty acid composition of Listeria monocytogenes nisin-resistant strains at 10°C and 30°C

Listeria monocytogenes isolates resistant to 10⁵ IU ml^-1 nisin were obtained at 30°C (NR30) and at 10°C (NR10). Nisin prolonged the lag phase of isolate NR30 at 10°C. Isolates NR30 and NR10 did not produce a nisinase. Protoplasts of isolate NR30 were unaffected by exposure to nisin. The fatty acid composition from the wild-type strain and NR isolates was determined. As expected, temperature-induced differences in the C15/C17 fatty acid ratios were found. Growth of the NR strains in the presence of nisin resulted in significantly different C15/C17 ratios and a significant increase in the percentage of C16:0, C16: 1, C18:0 and C18: 1 fatty acids at 10°C and 30°C. Both the NR10 and NR30 isolates had similar growth rates at low temperatures, but these were slower than the wild-type strain. These results indicate that 'nisin resistance'is an environmentally defined phenotype and that nisin induces changes in the fatty acid composition of the membrane in L. monocytogenes nisin-resistant isolates regardless of the growth temperature.     

Methanogenesis and methanotrophy within a Sphagnum peatland

Abstract: Methane production and consumption activities were examined in a Massachusetts peatland. Peat from depths of 5–35 cm incubated under anaerobic conditions, produced an average of 2 nmol CH₄ g⁻¹ h⁻¹ with highest rates for peat fractions between 25–30 cm depth. Extracted microbial nucleic acids showed the strongest relative hybridization with a 16S rRNA oligonucleotide probe specific for Archaea with samples from the 25–30 cm depth. In aerobic laboratory incubations, the peat consumed methane with a maximum velocity of 67 nmol CH₄ g⁻¹ h⁻¹ and a K _s of 1.6 μM. Methane consumption activity was concentrated 4–9 cm below the peat surface, which corresponds to the aerobic, partially decomposed region in this peatland. Phospholipid fatty acid analysis of peat fractions demonstrated an abundance of methanotrophic bacteria within the region of methane consumption activity. Increases in temperature up to 30°C produced an increase in methane consumption rates for shallow samples, but not for samples taken from depths greater than 9 cm. Nitrogen fixation experiments were carried out using ¹⁵N₂ uptake in order to avoid problems associated with inhibition of methanotrophy. These experiments demonstrated that methane in peat samples did not stimulate nitrogen fixation activity, nor could activity be correlated with the presence of methanotrophic bacteria in peat fractions.     

Starch utilization by the human large intestinal microflora

High levels (2–565 units/g) of amylase activity were observed in human faeces. Over 92% of amylase activity in faeces obtained from healthy persons was extracellular, whereas only about 9% of activity was associated with particulate material and washed cells. Bacterial cell-bound amylases were considerably more efficient in breaking down starch, however, than were the soluble enzymes which occurred in cell-free faecal supernatant fluids. Cell population densities of anaerobic starch-hydrolysing bacteria in the stools of ten persons ranged from 1.1 × 10¹⁰ to 3.3 × 10¹²/g of faeces. Identification of 120 starch-hydrolysing colonies isolated from the stools of six subjects showed that the predominant amylolytic bacteria belonged to the genera Bifidobacterium, Bacteroides, Fusobacterium and Butyrivibrio. Mixed populations of gut bacteria rapidly fermented starch with the production of volatile fatty acids and organic acids. Lactate was observed to be a major, though transient intermediate during starch fermentation by these cultures. Approximately 60% of starch utilized was converted to volatile fatty acids, which in the human colon would be potentially available for absorption.     

Relation between Aeromonas and faecal coliforms in fresh waters

A possible correlation between the presence of mesophilic aeromonads and the number of faecal coliforms present in three fresh-water habitats subject to differing levels of faecal pollution was investigated. Concentration of Aeromonas spp. between 10²–10⁹ cfu/100 ml and faecal coliforms of between 9–10⁷ cfu/100 ml were found in the waters. In water free from faecal pollution there was no correlation but in polluted waters there was a significant relationship between the numbers of aeromonads, faecal coliform and the concentration of organic matter measured by biological oxygen demand.     

Genetic and environmental influences on self-reported and parent-reported behavior problems in young adult adoptees

The aim of the present study was to estimate the genetic, shared and nonshared environmental contributions to self-reported and parent-reported internalizing and externalizing problems in a follow-up study of intercountry adopted young adults. Young Adult Self-Report ratings were obtained from 1475 adoptees aged 22–32 years and Young Adult Behavior Checklist ratings from 1115 adoptive parents. For the genetic analyses, a subset of 143 adopted biologically related and 295 unrelated siblings was used. The data were subjected to model fitting decomposing three sources of variance: genetic factors (A) shared environment (C) and nonshared environment (E). Genetic factors were of more importance in both self-reported ( A ²= 54%, C ²= 0, and E ²= 46%) and parent-reported ( A ²= 76%, C ²= 15% and E ²= 9%) internalizing problems. Environmental factors were of more importance in both self-reported ( A ²= 33%, C ²= 17% and E ²= 50%) and parent-reported ( A ²= 28%, C ²= 27% and E ²= 45%) externalizing problems. This was in contrast with findings from the first and second assessments in the same sample during adolescence when genetic factors were more important in explaining externalizing problems compared with internalizing problems. Our results suggest a developmental reversal in genetic and environmental influences on behavior problems from early adolescence into adulthood, which could be related to different underlying developmental trajectories.     

Release of Neurotransmitter Amino Acids from Synaptosomes: Enhancement of Calcium-Independent Efflux by Oleic and Arachidonic Acids

Abstract: The release of preloaded [¹⁴C]neuroactive amino acids (glutamic acid, proline, γ-aminobutyric acid) from rat brain synaptosomes can occur via a time-dependent, Ca²⁺ -independent process. This Ca²⁺-independent efflux is increased by compounds that activate Na⁺ channels (veratridine, scorpion venoms), by the ionophore gramicidin D, and by low concentrations of unsaturated fatty acids (oleic acid and arachidonic acid). Saturated fatty acids have no effect on the efflux process. Neither saturated nor unsaturated fatty acids have an effect on the release of [¹⁴C]leucine, an amino acid not known to possess neurotransmitter properties. The increase in the efflux of neuroactive amino acids by oleic and arachidonic acids can also be demonstrated using synaptosomal membrane vesicles. Under conditions in which unsaturated free fatty acids enhance amino acid efflux, no effect on ²²Na⁺ permeability is observed. Since Na⁺ permeability is not altered by fatty acids, the synaptosomes are not depolarized in their presence and, thus, the Na⁺ gradient can be assumed to be undisturbed. We conclude that unsaturated fatty acids represent a potentially important class of endogenous modulators of neuroactive amino acid transport in nerve endings and further postulate that their action is the result of an uncoupling of amino acid transport from the synaptosomal Na⁺ gradient.     

Identification of Strychnine Binding Sites in the Rat Retina

Abstract: [³H]Strychnine specifically binds to membrane fractions isolated from rat retinae. The binding is saturable, with an apparent dissociation constant, K _D, of 14.3 × 10⁻⁹ M and 205 fmol bound/mg protein. Specific binding is time-dependent and proportional to protein concentration. Glycine and taurine are equally potent inhibitors of [³H]strychnine binding ( K _i= 4 × 10⁻⁵ M); no other amino acids endogenously present in the retina inhibited [³H]strychnine binding.     

An SNP in the goat CSN2 promoter region is associated with the absence of beta-casein in milk

So far, at least eight alleles in the goat CSN2 locus have been associated with the level of β -casein expression in milk. Alleles CSN2 ^A , CSN2 ^{A 1}, CSN2 ^B , CSN2 ^C , CSN2 ^D and CSN2 ^E have been associated with normal content (allele effects of about 5 g of β -casein per litre), whereas the CSN2 ⁰ and CSN2 ⁰¹ alleles have been associated with non-detectable levels of β -casein. Most of these alleles have been characterized genetically. Herein, we report the identification of a previously unreported SNP in the goat CSN2 promoter region ( AJ011018 :g.1311T>C), which is associated with the absence of β -casein in the milk. Furthermore, we developed a PCR-based method that allows detection of this mutation.     

Quantification of pathogenic micro-organisms in the sludge from treated hospital wastewater

The sludge from hospital waste treatment facilities is a potential source of infectious organisms. The average numbers of micro-organisms in the sludge of hospital wastewater in Taiwan were as follows: total count 8·1 × 10⁷ cfu g⁻¹ (dry weight of sludge), and 1·4 × 10⁶, 3·6 × 10⁵, 1·6 × 10⁵, 2·2 × 10⁵ and 5·5 × 10⁴ cfu g⁻¹ (dry weight of sludge) for total coliforms, faecal coliforms, faecal streptococci, Pseudomonas aeruginosa and Salmonella spp., respectively . Salmonella spp. were detected in 37% (10 of 27) of the sludges from hospital wastewaters. Therefore, the treatment of such sludge to reduce pathogenic micro-organisms should be considered.     

Physiological mechanisms of buoyancy in eggs from brackish water cod

Newly fertilized eggs of brackish water (Gotland, Baltic Sea) and marine (Lofoten, Norway) cod were investigated with regard to specific gravity, wet and dry weight, water content, chorion weight, and content of protein, free amino acids (FAA), and ions. The eggs had neutral buoyancies equivalent to a salinity of 14^.3% (range 11^.5–16^.2%) in brackish water, and 33^.0% (range 31^.8–34^.5%) in the marine environment. A buoyancy model was developed and showed that this difference was mainly caused by differences in egg water content which was 96.6 ± 0^.47% and 92.7 ± 0.45% in the brackish and marine eggs, respectively. The higher water content of the brackish eggs resulted from increased water uptake during final oocyte maturation due to higher intracellular contents of FAA, Cl ^– and NH₄⁺. SDS polyacrylamide gel electrophoresis of eggs and oocytes, and measurements of egg protein content suggested that the FAA pool of both egg types originated from hydrolysis of specific yolk proteins. The main contributor seemed to be a protein with a molecular weight of 100 kDa.     

Utilization of Uniformly Labeled 13C-Polyunsaturated Fatty Acids in the Synthesis of Long-Chain Fatty Acids and Cholesterol Accumulating in the Neonatal Rat Brain

Abstract: Polyunsaturated fatty acids are needed for normal neonatal brain development, but the degree of conversion of the 18-carbon polyunsaturated fatty acid precursors consumed in the diet to their respective 20-and 22-carbon polyunsaturates accumulating in the brain is not well known. In the present study, in vivo ¹³C nuclear magnetic resonance spectroscopy was used to monitor noninvasively the brain uptake and metabolism of a mixture of uniformly ¹³C-enriched 16-and 18-carbon polyunsaturated fatty acid methyl esters injected intragastrically into neonatal rats. In vivo NMR spectra of the rat brain at postnatal days 10 and 17 had larger fatty acid signals than in uninjected controls, but changes in levels of individual fatty acids could not be distinguished. One day after injection of the U-¹³C-polyunsaturated fatty acid mixture, ¹³C enrichment (measured by isotope ratio mass spectrometry) was similar in brain phospholipids, free fatty acids, free cholesterol, and brain aqueous extract; ¹³C enrichment remained high in the phospholipids and cholesterol for 15 days. ¹³C enrichment was similar in the main fatty acids of the brain within 1 day of injection but 15 days later had declined in all except arachidonic acid while continuing to increase in docosahexaenoic acid. These changes in ¹³C enrichment in brain fatty acids paralleled the developmental changes in brain fatty acid composition. We conclude that, in the neonatal rat brain, dietary 16-and 18-carbon polyunsaturates are not only elongated and desaturated but are also utilized for de novo synthesis of long-chain saturated and monounsaturated fatty acids and cholesterol.     

Efflux of ions and ATP depletion induced by pediocin PA-1 are concomitant with cell death in Listeria monocytogenes Scott A

Domination of Carnobacterium divergens LV13 by a bacteriocin-producing (bac⁺) organism Carnobacterium piscicola LV17 was dependent on the level of inoculum of the producer strain and its bacteriocin production. When C. piscicola LV17 was grown in APT broth from an initial inoculum of α-10⁴ cfu ml^-1, bacteriocin was not produced (bac^-) although maximum population was reached. The culture remained bac^- during subsequent inoculation at 10²-10⁷ cfu ml^-1 unless it was first grown on solid medium or if heat-treated supernatant fluids from a bac⁺ culture of C. piscicola LV17, LV17A or LV17B were added to the culture prior to the stationary phase of growth. Use of purified carnobacteriocins from C. piscicola LV17A and LV17B confirmed their role in regulation of the bac⁺ phenotype. The need for induction might account in part for differences in bacteriocin production by cultures in liquid and on solid growth media.     

Carbon isotope-labelling experiments indicate that ladderane lipids of anammox bacteria are synthesized by a previously undescribed, novel pathway

Ladderane lipids are unusual membrane lipids of bacteria that anaerobically oxidize ammonium to dinitrogen gas (anammox). Ladderane lipids contain linearly concatenated cyclobutane rings for which the pathway of biosynthesis is currently unknown. To investigate the possible biosynthetic routes of these lipids, 2-¹³C-labelled acetate was added to a culture of the anammox bacterium Candidatus Brocadia fulgida. Labelling patterns obtained by high-field ¹³C nuclear magnetic resonance spectroscopy of isolated lipids indicated that C . Brocadia fulgida synthesizes C_16:0 and iso C_16:0 fatty acids according to the known pathway of type II fatty acid biosynthesis. The ¹³C-labelling pattern of the C₈ alkyl chain of the C₂₀ [3] ladderane monoether also indicated the use of this route. However, carbon atoms in the cyclobutane rings and the cyclohexane ring were nonspecifically labelled and did not correspond to known patterns of fatty acid synthesis. Taken together, our results indicate that it is unlikely that ladderane lipids are formed from the cyclization of polyunsaturated fatty acids as hypothesized previously and suggest an alternative, although as yet unknown, pathway of biosynthesis.     

Improvement in the growth performance of white shrimp, Litopenaeus vannamei, by a protease-producing probiotic, Bacillus subtilis E20, from natto

Aims:  To isolate and identify a benefic bacterium, Bacillus subtilis E20, from natto (fermented soybeans), and incorporate it into shrimp feed to promote shrimp growth performance.
Methods and Results:  A protease-producing bacterium, E20, isolated from natto was identified as B. subtilis by an API 50 CHB kit and the 16S rDNA sequence. B. subtilis E20 was able to grow at a broad range of temperatures (10–50°C), pH values (5–10), and NaCl levels (0–9%). The best culture conditions for B. subtilis E20 to produce the protease were 40°C, a pH of 6–8 and 0% NaCl. No shrimp died after being injected with B. subtilis E20 [up to 10⁹ colony-forming units (CFU) per shrimp]. Bacillus subtilis E20 was incorporated in diets at the levels of 0 (control), 10⁶, 10⁷, and 10⁸ CFU kg⁻¹ for shrimp grow-out culture, and results showed that after feeding on B. subtilis E20-containing diets (10⁸ CFU kg⁻¹ of diet), shrimp had excellent growth performance and production compared to the control because protease activities in the digestive tract were improved by B. subtilis E20.
Conclusions:  Bacillus subtilis E20 isolated from natto is a great protease producer and is able to improve shrimp growth performance through increasing the digestibility of food.
Significance and Impact of the Study:  Results suggest that B. subtilis E20 is a potential candidate for use as a probiotic to improve shrimp growth performance, and consequently reduce feed costs.     

Description of Oxalicibacterium horti sp. nov. and Oxalicibacterium faecigallinarum sp. nov., new aerobic, yellow-pigmented, oxalotrophic bacteria

Three strains of aerobic, Gram-negative, rod-shaped, non-spore-forming, yellow-pigmented bacteria (OD1^T, YOx^T and NS13), which were isolated in previous studies by enrichment in a mineral medium with potassium oxalate as the sole carbon source, were characterized. On the basis of 16S rRNA gene sequence similarity, strains OD1^T, YOx^T and NS13 belong to the Betaproteobacteria , most closely related to Oxalicibacterium flavum TA17^T (97.2–99.7% sequence similarity). The major whole-cell fatty acids were C_16:0, C_16:1ω7c and C_17:0 cyclo. The results of DNA–DNA hybridization and physiological and biochemical tests allowed genotypic and phenotypic differentiation of strains OD1^T and YOx^T from O. flavum TA17^T and from each other. Therefore, it is concluded that the strains OD1^T and YOx^T represent novel species within the genus Oxalicibacterium , for which the names Oxalicibacterium horti sp. nov. (type strain OD1^T=DSM 21640^T=NBRC 13594^T) and Oxalicibacterium faecigallinarum sp. nov. (type strain YOx^T=DSM 21641^T=CCM 2767^T) are proposed.     

Fatty acid composition and microbial activity of benthic marine sediment from McMurdo Sound, Antarctica

Abstract Signature lipids from the phospholipid esterlinked fatty acids (PELFA) of cell membranes were used to describe benthic microbial communities of 4 Antarctic sediments. Metabolic activities of the communities were determined by incorporation of [³H]thymidine into bacterial DNA and sodium [¹⁴C]acetate into membrane lipids. Biomass measurements from extractable phospholipid fatty acids per g dry wt. ranged between 6 to 76 nmol, or when converted to number of bacteria, 3.7 × 10⁸ to 4.5 × 10⁹ cells per g dry wt. The West Sound site at New Harbor contained the lowest biomass, while Cape Evans on the East Sound contained the greatest. A marked difference was also noted between sites in their sediment microbial community structure. The East Sound sites at Cape Armitage and Cape Evans contained a greater abundance of diatom marker lipids, whilst both sides of the Sound contained approximately the same relative amounts of bacterial groups distinguished using PELFA. Activity of sediment microorganisms measured by radiolabel incorporation under ambient conditions followed the trends of the biomass measurements. The East Sound sites were more active by an average of 45–73% for [³H]thymidine and possibly also for sodium [¹⁴C]acetate.