V. Boote

Phospholipid analogues of Porphyromonas gingivalis

Porphyromonas has lipids containing hydroxy acids and C16:0 and iso‐C15:0 major monocarboxylic acids among others. Nothing is known of its individual phospholipid molecular species. The aim of this study was to determine molecular weights and putative identities of individual phospholipid molecular species extracted from Porphyromonas gingivalis (seven strains), P. asaccharolytica (one strain) and P. endodontalis (two strains). Cultures on Blood‐Fastidious Anaerobe Agar were harvested, washed and freeze‐dried. Phospholipids were extracted and separated by fast atom bombardment mass spectrometry (FAB MS) in negative‐ion mode. Phospholipid classes were also separated by thin layer chromatography (TLC). The major anions in the range m/z 209–299 were consistent with the presence of the C13 : 0, C15 : 0, C16 : 0 and C18 : 3 mono‐carboxylate anions. Major polar lipid anion peaks in the range m/z 618–961 were consistent with the presence of molecular species of phosphatidylethanolamine, phosphatidylglycerol and with unidentified lipid analogues. Porphyromonas gingivalis differed from comparison strains of other species by having major anions with m/z 932, 946 and 960. Unusually, a feline strain of P. gingivalis had a major peak of m/z 736. Selected anions were studied by tandem FAB MS which revealed that peaks with m/z 653 and 946 did not correspond to commonly occurring classes of polar lipids. They were however, glycerophosphates. It is concluded that the polar lipid analogue profiles obtained with Porphyromonas are quite different from those of the genera Prevotella and Bacteroides but reveal heterogeneity within P. gingivalis.

Phospholipid molecular species distribution of some medically important Candida species analysed by fast atom bombardment mass spectroscopy

The aim of this study was to obtain detailed information on phospholipids (PL) of the medically important Candida species and to determine their possible chemotaxonomic significance. Lipids were extracted from 22 strains representing 8 Candida species and their PL molecular species distributions were determined by Fast Atom Bombardment Mass Spectroscopy (FAB MS) in negative ion mode. Fifteen major lower mass peaks (m/z 221 to 289) were attributable to the expected presence of carboxylate anions and 24 major higher mass peaks (m/z 557 to 837) were attributable to phospholipid anions. Major carboxylate peaks were of the following m/z and identities : 253, C16:1 ; 255, C16:0 ; 277, C18:3 ; 279, C18:2 ; 281, C18:1 ; and 283, C18:0. The most abundant peaks consistent with the presence of phospholipid molecular species anions include those of m/z 673, 743, 833, 834 and 836 tentatively identified as phosphatidic acid (PA) (34 : 1), phosphatidylglycerol (PG) (34 : 3), phosphtidylinositol (PI) (34 : 2) and two unknown molecular species. This profile is diagnostic for the genus Candida. Quantitative differences were observed between different Candida species. Thus, polar lipid molecular species distribution in Candida spp. has chemotaxonomic significance, especially so in the case of carboxylate anions.

Comparative phospholipid analogue distributions of Porphyromonas gingivalis isolated from cats in Australia and the USA.

DNA-DNA homology measurements and phospholipid (PL) analogue profiling have shown heterogeneity of Porphyromonas gingivalis. The aim of this study was to determine whether there were differences between cat strains of P. gingivalis from Australia and USA with respect to PL analogue distribution. Lipids were extracted with chloroform-methanol and examined by fast atom bombardment-mass spectrometry (FAB-MS) in negative-ion mode, using published methods. For PL analogues, the major anions included those with mass-to-charge (m/z)=634, 648, 662, 705, 932, 946 and 960, respectively, corresponding to expected presence of PE (28:0), PE (29:0), PE (30:0), PG (32:1), and three unknown homologues of a glycero-phospholipid with a single nitrogen. Analyses were compared to calculate a matrix of Pearson coefficients of linear correlation from which a dendrogram was produced of strains clustered by single linkage. One cluster was comprised solely of Australian cat-to-cat bite isolates and a second cluster included exclusively USA cat- and dog-to-human bite isolates except for one Australian cat-to-cat bite isolate (VPB 5089). The US cluster included three outliers, one of which was the Australian cat isolate VPB 5089. The human type strain (ATCC 33277) was quite remote from all dog and cat strains. It was shown that P. gingivalis human and non-human animal isolates have distinct PL analogue profiles from each other. Furthermore, the cat strains from the USA and those from Australia showed quantitative differences in polar lipid profiles that correlated largely with country of isolation.

Characterization of polar lipids of oral isolates of Candida, Pichia and Saccharomyces by Fast Atom Bombardment Mass Spectrometry (FAB MS).

Aims: To characterize fatty acid and phospholipid analogue profiles of oral yeasts.

Phospholipid molecular species distribution of Porphyromonas asaccharolytica ATCC 25260T: effects of temperature, culture age and pH

A.M. TAVANA, D.B. DRUCKER AND V. BOOTE. 1998. The aim of this study was to determine the extent to which phospholipid molecular species profiles are affected by different environmental factors in Porphyromonas asaccharolytica ATCC 25260T. Phospholipids were analysed by Fast Atom Bombardment Mass Spectrometry (FAB‐MS) in negative‐ion mode. Under standard growth conditions (37°C, pH7·0, 48 h), the most intense high mass anions were m/z 653 and 662. The latter is consistent with the expected presence of PE (30:0). The only changes in profiles were quantitative. These were compared using the Pearson Coefficient of Linear Correlation. The r‐values for initial pH comparisons ranged from 0·82 (pH 7·0 vs pH 6·0) to 1·00 (pH 5·0 vs pH 8·0), for incubation period, from 0·86 (48 vs 72 h) to 0·97 (96 vs 168 h), and for temperature, from 0·57 (40 vs 37°C) to 0·96 (37 vs 36°C). Differences were also seen when plates were incubated in anaerobe jars as opposed to an anaerobic work station (r = 0·75). It is concluded that it is essential to standardize growth parameters, and to use an anaerobe jar or an anaerobe work station, but not both.

Phospholipid molecular species distributions of Candida isolates from the UK and Iran

Aims: Some species of Candida have been shown to differ with respect to their polar lipid fingerprints when analysed by fast atom bombardment mass spectrometry (FABMS). The aims of this study were to contribute to the existing body of information by (i) examining representatives of species not previously examined and (ii) seeking strains differences associated with country of origin (UK or Iran).