The ability of Listeria monocytogenes to adapt to low temperature, low pH, and osmotic stress, to withstand cleaning and disinfection agents, to induce stress-related cross-protection responses, and to form biofilms, enables it to prevail and survive most of the common safety measures employed in industrial-scale food processing to ensure food safety. The objective of this study was to determine the changes that occur in the lipid composition of the membrane of L. monocytogenes under the conditions of the manufacture and storage of a laboratory-prepared soft cheese (feta type). The results show that L. monocytogenes cells present in the raw milk may survive the cheese manufacturing process as well as the storage period and that the history of the inoculated cells has an effect on their behavior during storage of the cheese. The bacterium seems to follow different pathways and adopt different strategies to preserve the optimal fluidity and functions of the membrane depending on the length of storage time and the history of the inoculated cells. In this context, cells derived from pre-inoculated cells grown in optimal or acidic conditions showed an increase in the number of straight chain fatty acids (SCFA) relative to branched ones (BCFA), without altering the ratio of BCFA C15:0 to BCFA C17:0 and the ratio of iso to anteiso FA. Alternatively, those cells derived from pre-inoculated cells grown in presence of benzalkonium chloride followed a different pathway. These cells achieved a balance between optimal fluidity and solid phase membrane not only by changes in SCFA content, but also by changes in the ratios of BCFA C15:0 to C17:0 and of iso to anteiso fatty acids. This latter approach to maintain membrane fluidity seems to be paired with the different survival trends of the bacterium under the conditions of this study.

This study addresses a major issue in microbial food safety, the elucidation of correlations between acid stress and changes in membrane fluidity of the pathogen Listeria monocytogenes. In order to assess the possible role that membrane fluidity changes play in L. monocytogenes tolerance to antimicrobial acids (acetic, lactic, hydrochloric acid at low pH or benzoic acid at neutral pH), the growth of the bacterium and the gel-to-liquid crystalline transition temperature point (T-m) of cellular lipids of each adapted culture was measured and compared with unexposed cells. The T-m of extracted lipids was measured by differential scanning calorimetry. A trend of increasing T-m values but not of equal extent was observed upon acid tolerance for all samples and this increase is not directly proportional to each acid antibacterial action. The smallest increase in T-m value was observed in the presence of lactic acid, which presented the highest antibacterial action. In the presence of acids with high antibacterial action such as acetic, hydrochloric acid or low antibacterial action such as benzoic acid, increased T-m values were measured. The T-m changes of lipids were also correlated with our previous data about fatty acid changes to acid adaptation. The results imply that the fatty acid changes are not the sole adaptation mechanism for decreased membrane fluidity (increased T-m. Therefore, this study indicates the importance of conducting an in-depth structural study on how acids commonly used in food systems affect the composition of individual cellular membrane lipid molecules.

This is the first study to evaluate the survival potential of cold-adapted Listeria monocytogenes in ice-cream. Cold adaptation enhances survival of this pathogen in ice-cream during the first period of storage compared to non-adapted cells. The viable population of cold-adapted and non-adapted cells was 3 log (36 days) and 4.3 log (27 days), respectively, lower than the initial population (6.3 log) in inoculated ice-cream. This behavior raises concerns for food safety. The viable population of both cold- and non-adapted cells displayed a slight statistical difference in the next period of frozen storage (0.29 and 0.75 log decline at 137 and 182 days, respectively). Significant numbers of L. monocytogenes cells survived for extended periods of time, irrespective of whether they were previously cold- or non-adapted (332 and 182 days respectively). The natural additives utilized (fructose syrup, corn syrup, sesame oil and sesame paste) did not have any significant effect on the response of non-adapted L. monocytogenes in ice-cream during 182 days of storage. On the other hand, the survival of cold-adapted L. monocytogenes is influenced by the ingredients utilized in the ice cream. Sesame paste and corn syrup had an inhibitory action on cold-adapted L. monocytogenes throughout the frozen storage (332 days) possibly as a consequence of lower water activity in samples with these additives.

Aims:This study provides a first approach to observing the alterations of the cell membrane lipids in the adaptation response of Listeria monocytogenes to the sanitizer benzalkonium chloride.Methods and Results:A thorough investigation of the composition of polar and neutral lipids from L. monocytogenes grown when exposed to benzalkonium chloride is compared to cells optimally grown. The adaptation mechanism of L. monocytogenes in the presence of benzalkonium chloride caused (i) an increase in saturated-chain fatty acids (mainly C-16:0 and C-18:0) and unsaturated fatty acids (mainly C-16:1 and C-18:1) at the expense of branched-chain fatty acids (mainly Ca-15:0 and Ca-17:0) mainly because of neutral fatty acids; (ii) no alteration in the percentage of neutral and polar lipid content among total lipids; (iii) a decrease in lipid phosphorus and (iv) an obvious increase in the anionic phospholipids and a decrease in the amphiphilic phosphoaminolipid.Conclusions:These lipid changes could lead to decreased membrane fluidity and also to modifications of physicochemical properties of cell surface and thus changes in bacterial adhesion to abiotic surfaces.Significance and Impact of the Study:The adaptation and resistance of L. monocytogenes to disinfectants is able to change its physiology to allow growth in food-processing plants. Understanding microbial stress response mechanisms would improve the effective use of disinfectants.

This study provides a first approach to observe the effects on Listeria monocytogenes of cellular exposure to acid stress at low or neutral pH, notably how phospho- or neutral lipids are involved in this mechanism, besides the fatty acid profile alteration. A thorough investigation of the composition of polar and neutral lipids from L. monocytogenes grown at pH 5.5 in presence of hydrochloric, acetic and lactic acids, or at neutral pH 7.3 in presence of benzoic acid, is described relative to cells grown in acid-free medium. The results showed that only low pH values enhance the antimicrobial activity of an acid. We suggest that, irrespective of pH, the acid adaptation response will lead to a similar alteration in fatty acid composition [decreasing the ratio of branched chain/saturated straight fatty acids of total lipids], mainly originating from the neutral lipid class of adapted cultures. Acid adaptation in L. monocytogenes was correlated with a decrease in total lipid phosphorus and, with the exception of cells adapted to benzoic acid, this change in the amount of phosphorus reflected a higher content of the neutral lipid class. Upon acetic or benzoic acid stress the lipid phosphorus proportion was analysed in the main phospholipids present: cardiolipin, phosphatidylglycerol, phosphoaminolipid and phosphatidylinositol. Interestingly only benzoic acid had a dramatic effect on the relative quantities of these four phospholipids.

Listeria monocytogenes membrane lipid fatty acids (FA) play a role in its cold adaptation. Our previous study indicated that in newly synthesized neutral lipids, from the early stage of the temperature response (3 days at 5 °C), only the anteiso-15:0 FA was selectively increased. In this study we focus on several NL class comparisons. Cells of L. monocytogenes were grown until stationary phase at 30 and 5 °C and the FA from 1.2 diglyceride (DG), 1.3 DG and free FA as well as NL (in bulk) from each culture were analysed by GC. At 30 °C, L. monocytogenes showed a FA profile from 1.2 DG, 1.3 DG, free FA and NL, dominated by anteiso-15:0 percentages 28.4%, 45.1%, 30.4%, 45.2%, respectively. Also from the three first classes the ratio anteiso-15:0/anteiso-17:0 FA averaged 2.0, but the ratio of branched chain FA to straight chain FA of each class was 1.2, 4.7 and 2.3, respectively. L. monocytogenes cold adaptation response was based singularly on the increase (×5.9, ×5.4, ×8.3 and ×7.3-fold) of the ratio anteiso-15:0/anteiso-17:0 FA for 1.2 DG, 1.3 DG, free FA and NL, respectively, in contrast to a negligible change of the ratio branched to straight chain FA. Using 1H-NMR analysis it was revealed that the % reduction of ratio variation [-CH2-]/[-CH3] at cold (4.65%) for the NL class of Listeria’s fatty acyl chains reflected the increase of the ratio value branched-15:0/branched-17:0 FA that was determined by GC analysis (×6.1-fold).

In this work a thorough consideration of the membrane lipid composition of Listeria monocytogenes together with DSC analysis is described in order to estimate the biological importance of lipid changes during low-temperature adaptation. Furthermore, these studies provide comparative data for fatty acid changes for neutral, NL and polar lipids, PL separately. The cold adaptation (5°C) response of L. monocytogenes showed (i) an increase in the level of NL content (30%) among the total lipids, TL and (ii) that the increase (7-fold) in the anteiso-15:0/anteiso-17:0 fatty acid ratio, FAr, for cold NL was at variance with the ratio for TL and PL (about 10-fold). We correlated our findings with DSC studies on phase transition temperature (Tc), enthalpy difference (ΔH) and peak range of the transition for TL, PL, NL (from cultures at 30 and 5°C); The decrease of Tc (10.5°C) and ΔH (51%) for TL is a reflection of the decrease of Tc (11.5°C) and ΔH (56%) for PL. This large decrease is interpreted by the high (10-fold) increase of a-15:0/a-17:0 FAr of PL5°C. In NL the decrease of Tc (3°C) and of ΔH (42%) is interpreted by both adaptation mechanisms: the (lower) 7-fold increase of anteiso-15:0/anteiso-17:0 FAr and the NL percentage calculated from increased mass values. The peak range of TL5°C (from −15 to 25°C) is a reflection of the peak range of NL5°C, which is unchanged, as is the peak range of NL30°C.

The goal of this study is to supplement the composition and nature of sphingophosphonolipids diversity from edible mollusks (Mytilus galloprovincialis, Eobania vermiculata) and from jellyfish Pelagia noctiluca, organisms rich in phosphonolipids. M. galloprovincialis contained a major ceramide 2-aminoethylphosphonate (CAEP-IM) and a minor ceramide that was detected chromatographically as the methyl analog (CAEP-IIM). In CAEP-IM, saturated fatty acids (FA) of 14, 16 and 18 carbons amounted to 68.8%; also 52.5% dihydroxy bases were detected. On thin layer chromatography, the Rf for CAEP-IIM was smaller than the Rf for CAEP-IM because of an increase of 22.0% in 2OH-16:0 FA, plus 29.2% trihydroxy bases (phytosphingosine). Similarly, a ceramide 2-methylaminoethylphosphonate (CAEP-IIE, 1.5% of phospholipids) was quantitated in Eobania (apart from the previously reported major CAEP, 7.6%). In CAEP-IIE, saturated and hydroxy FA of 14, 16 and 18 carbons amounted to 37.0 and 37.8%; 29.1% dihydroxy and 23.0% trihydroxy bases were detected in the same molecule. Eobania's unsaturated FA percentages (total lipids: 66.3, polar: 47.5, neutral: 59.0) were similar to those previously found for other land snails. A suite of two minor CAEP (CAEP-IIP, CAEP-IIIP) was quantitated in Pelagia at 2.0 and 1.3% of phospholipids (apart from the previously reported major CAEP, 21.0%) identified chromatographically as methyl analogs. In CAEP-IIP, saturated FA of 14, 16, 18 and 19 carbons amounted to 56.0%; 12.6% dihydroxy and 34.1% trihydroxy bases were also detected in CAEP-IIP. The Rf CAEP-IIIP<Rf CAEP-IIP owing to an increase of +8.5% of hydroxy FA and +12.3% of trihydroxy bases. The compositions of CAEP-IIM and CAEP-IIE appear to be specific of each organism, while the composition of molluscan or jellyfish major sphingophosphonolipids appears not specific.

The goal of this study is to elucidate and identify several sphingophosphonolipids from Aurelia aurita, an abundant but harmless Aegean jellyfish, in which they have not previously been described. Total lipids of A. aurita were 0.031–0.036% of fresh tissue, and the lipid phosphorus content was 1.3–1.7% of total lipids. Phosphonolipids were 21.7% of phospholipids and consisted of a major ceramide aminoethyl-phosphonate (CAEP-1; 18.3%), as well as three minor CAEP (II, III, IV) methyl analogs at 1.3, 1.1, and 1.0%, respectively. The remaining phospholipid composition was: phosphatidylcholine, 44.5%, including 36.2% glycerylethers; phosphatidylethanolamine, 18.6%, including 4.5% glycerylethers; cardiolipin, 5.6%; phosphatidylinositol, 2.6%; and lysophosphatidylcholine, 5.0%. In CAEP-1; saturated fatty acids of 14–18 carbon chain length were 70.8% and were combined with 57.3% dihydroxy bases and 23.4% trihydroxy bases. The suite of the three minor CAFP methyl analogs were of the same lipid class based on the head group, but they separated into three different components because of their polarity as follows: CAEP-II and CAEP-III differentiation from the major CAEP-I was mainly due to the increased fatty acid unsaturation and not to a different long-chain base, but the CAEP-IV differentiation from CAEP-I, apart from fatty acid unsaturation, was due to the increased content of hydroxyl groups originated from both hydroxy fatty acids and trihydroxy long-chain bases. Saturated fatty acids were predominant in total (76.7%), polar (83.0%), and neutral lipids (67.6%) of A. aurita. The major phospholipid components of A. aurita were comparable to those previously found in a related organism (Pelagia noctiluca), which can injure humans.

One of the possible roles of phosphonolipids is that they have a contribution to the protection of cellular integrity and survival of aquatic organisms (mollusca, cnidaria) as these lipids are included at high percentages. The total lipids of the edible mussel Mytilus galloprovincialis (Mollusca, Bivalvia, Mytilidae) were found to constitute 1.27% of fresh tissue. Polar lipid components constitute 61.5% of the total lipids. After separation by Solid Phase Extraction, the polar lipid fraction was separated by two dimensional thin-layer chromatography and the total phosphorus of each component, was determined. The main polar lipids found were: Phosphatidylcholine, 41.6 ± 0.8% (of which 11.3 ± 0.5% was glyceryl ether analog); ceramide aminoethylphosphonate 11.2 ± 0.2% plus 2.8 ± 0.1% another minor species; phosphatidylethanolamine, 26.6 ± 0.5% (of which 12.2 ± 0.3% was glyceryl ether analog). The individual ceramide aminoethylphosphonate species were isolated by preparative thin layer chromatography and the structure of the major one was confirmed by a combination of analytical and chromatographic methods. Saturated fatty acyl groups with 16 carbon atoms were the main components (48.4%) of the major ceramide aminoethylphosphonate species. Diglyceride aminoethyl phosphonates were not found in lipids of M. galloprovincialis.

Listeria monocytogenes, a Gram-positive pathogenic food-borne bacterium, is notably resistant to chill and osmotic stress.Listeriais characterized by an unusual proportion of branched-chain fatty acid (>85%), primarily anteiso-15:0 and anteiso-17:0. Cells grown at lower temperatures contain significantly less anteiso-17:0 than those grown at higher temperature. Cold-shock studies were performed to determine the mechanisms by whichListeriaperceives cold shock and changes its membrane composition. This study made three comparisons: (i) The lipids ofL. monocytogenescultured at 30°C were compared with those of cells grown at 6°C. Extraction of total lipids yielded 4–4.5 and 6.5–7 mg ml-1cell mass (wet weight), respectively. Total lipids contained 3–4% and 2–2.5% lipid phosphorus, respectively, reflecting a higher content of neutral lipids in cells grown at 6°C. (ii) The fatty-acid compositions of different lipid classes (total lipids, the major phospholipid, and the glyco-containing phospholipid class) all responded by increasing the proportion of anteiso-15:0 fatty acids at low temperature. (iii) The fatty-acid composition of the neutral and polar lipids from the early stage of the temperature response (3 days at 5°C) and the effect of chloramphenicol or cerulenin were examined. The results indicated that anteiso-15:0 fatty acids were selectively increased in the newly synthesized neutral lipids, but the polar lipids did not show an apparent change in fatty-acid composition. These changes in membrane fatty-acid composition did not require new protein synthesis (the process was chloramphenicol insensitive) and the existing condensing enzyme activity was necessary to the fatty-acid response (the process was cerulenin sensitive).

Listeria monocytogenes is a Gram-positive bacterium that can adapt to high salinity and cold. Because the membrane lipids may play a role in its survival and adaptation, we have examined the polar lipids of L. monocytogenes. Extraction of total lipids from L. monocytogenes yielded 7 +/- 1 mg/mL wet cells. Polar lipids represented 64% of total lipids and contained 9% lipid-phosphorus. Polar lipids were separated into 14 components by two-dimensional thin-layer chromatography. Eight components (88% of polar lipids) contained lipid-phosphorus; among these was one major component (34% of polar lipids). Two other phospholipids were ninhydrin-positive components and accounted for 15% of the polar lipids. Orcinol staining revealed two glyco- or sulfo-lipids accounting for 9% of polar lipids. Five components (4% of polar lipids) were amino components free of phosphorus. The major component contained 46% of its fatty acids as 15:0 anteiso, 24% as 17: 0 anteiso, and 11% as 15:0 iso. The fatty acid profile of the remaining polar lipids was variable, consisting primarily of 16:0, 18:0, 15:0 anteiso, and 17:0 anteiso. Their unsaturation level was < or = 20%; however, the major phosphoaminolipid component was 46% unsaturated. The ratios of 15:0 anteiso/17:0 anteiso and 15:0 anteiso/15:0 iso were similar in all classes, averaging 1.5 and 4.5, respectively. Since the adaptation process to stressful environments involves activation of a membrane transport system for the protectant glycine betaine, the membrane lipids may play a role in enabling transport.

Listeria monocytogenes is a Gram-positive bacterium that causes meningitis, septicemia and death in humans. Found in low-acid cheeses, vegetables and meat, L. monocytogenes is resistant to osmotic and chill stress. Food handling practices that suppress microbial competitors can therefore promote its growth. In response to hyperosmotic or chill stress, L. monocytogenes accumulates the potent protectant glycine betaine from the medium, which decreases the lag time and increases the growth rate of the organism. The molecular basis for activation of glycine betaine transport by chill (7 °C), despite the expected membrane lipid phase transition, may reside in the lipid composition. The present research identified the lipids of L. monocytogenes. Extraction of total lipids yielded 7 ± 1 mg ml−1 wet cells, with a 5–6% phosphorus content. Polar lipids represented 64% of total lipids. There was a clear difference in the relative complexity of the fatty acids: neutral lipids were more varied and unsaturated fatty acids represented 19% of the total. Polar lipid fatty acids were primarily 15:anteiso (50%) and 17:anteiso (25%).

The constituent lipids of Greek Honey have been isolated and studied by an initial simple extraction procedure (comparable to that of counter-current distribution) and consequent chromatographic separation on a silicic acid column. The fractions collected were subjected to: (I) qualitative T.L.C before or after saponification and (II) gas-chromatographic analysis of methyl esters. In addition to the fatty acids mentioned by other investigators, honey has been found to contain a number of neutral lipids, albeit in small amounts, i.e. hydrocarbons, waxes, cholesterol esters, fatty acid esters, fatty acids, fatty alcohols, sterols, dihydroxy and trihydroxy compounds, as well as some esters of polyols. The probable nature of another three unknown constituents is also discussed.