Biomedical subjects

Tania C Sorrell

Publications and source records attributed to Tania C Sorrell.

26 records · Page 2Linked to original sources

PubMed2003

Identification of metabolites of importance in the pathogenesis of pulmonary cryptococcoma using nuclear magnetic resonance spectroscopy.

Primary lung infection with Cryptococcus neoformans is characterised by circumscribed lesions (cryptococcomas). To identify cryptococcal and/or host products of importance in pathogenesis, we applied proton nuclear magnetic resonance (NMR) spectroscopy, which identifies mobile compounds present in complex mixtures, to experimental pulmonary cryptococcomas from rats. Magnetic resonance experiments were performed on cryptococcomas (n = 10) and healthy lungs (n = 8). Signal assignment to key metabolites was confirmed by homo-nuclear and hetero-nuclear NMR correlation spectroscopy. Cryptococcal metabolites, dominating spectra from cryptococcomas included the stress protectants, trehalose and mannitol, acetate, and in some animals, ethanol. Glycerophosphorylcholine was also abundant in cryptococcomas, consistent with hydrolysis of phospholipids in vivo by the cryptococcal enzyme, phospholipase B (PLB). PLB has been identified by molecular studies as a cryptococcal virulence determinant. We propose that PLB secreted by cryptococci promotes tissue invasion by hydrolysing host phospholipids, such as dipalmitoyl phosphatidyl choline, which is abundant in pulmonary surfactant, and lung cell membrane phospholipids. Our results confirm the utility of NMR spectroscopy in studies of microbial pathogenesis.

Animals↗

PubMed2003

PCR-restriction fragment length polymorphism analysis of the phospholipase B (PLB1) gene for subtyping of Cryptococcus neoformans isolates.

Cryptococcus neoformans is a pathogenic yeast that is currently divided into three varieties, five serotypes, and eight molecular types. The following report describes the use of PCR-restriction fragment length polymorphism (RFLP) analysis of the phospholipase B gene (PLB1) as a simple tool to differentiate between C. neoformans subgroups. A PLB1 fragment, 1,970 bp, was amplified and digested with either AvaI or HindIII. Both sets of profiles grouped the isolates into their respective varieties, but only the AvaI profiles allowed for the identification of the eight molecular types via the corresponding RFLP profiles A1 to A8. Digestion of the same fragments with HindIII resulted in RFLP profiles H1 to H5, which distinguished only between serotype A, AD, D, and B/C. Neither enzyme distinguished serotype B from serotype C. The serotype AD profile was a composite of the serotype A and D profiles. Further investigation showed that the serotype AD isolates used in this study are heterozygous, with one allele of PLB1 originating from a serotype A parent and the other from a serotype D parent.

Cryptococcus neoformans↗

PubMed2003

Rapid identification of Candida species by using nuclear magnetic resonance spectroscopy and a statistical classification strategy.

Nuclear magnetic resonance (NMR) spectra were acquired from suspensions of clinically important yeast species of the genus Candida to characterize the relationship between metabolite profiles and species identification. Major metabolites were identified by using two-dimensional correlation NMR spectroscopy. One-dimensional proton NMR spectra were analyzed by using a staged statistical classification strategy. Analysis of NMR spectra from 442 isolates of Candida albicans, C. glabrata, C. krusei, C. parapsilosis, and C. tropicalis resulted in rapid, accurate identification when compared with conventional and DNA-based identification. Spectral regions used for the classification of the five yeast species revealed species-specific differences in relative amounts of lipids, trehalose, polyols, and other metabolites. Isolates of C. parapsilosis and C. glabrata with unusual PCR fingerprinting patterns also generated atypical NMR spectra, suggesting the possibility of intraspecies discontinuity. We conclude that NMR spectroscopy combined with a statistical classification strategy is a rapid, nondestructive, and potentially valuable method for identification and chemotaxonomic characterization that may be broadly applicable to fungi and other microorganisms.

Candida↗

PubMed2002

Strain-dependent effects of environmental signals on the production of extracellular phospholipase by Cryptococcus neoformans.

Extracellular phospholipase (PL) activities comprising phospholipase B, lysophospholipase and lysophospholipase transacylase have been identified in culture supernatants of Cryptococcus neoformans and contribute to virulence. We found that PL production was optimal after fungal growth at 30 degrees C and secretion at 37 degrees C for all six C. neoformans isolates studied (four C. neoformans var. neoformans and two C. neoformans var. gattii). No increase in PL activity was found in one strain, NU-2, in low iron or tissue culture media, conditions where upregulation of other virulence factors has been reported. The most virulent strains in an intravenous mouse model of infection were best able to produce PL at growth and secretion temperatures of 37 degrees C, in tissue culture media and under assay conditions of pH 7.0.

Buffers↗

PubMed2002

Cerebral nocardiosis characterized by magnetic resonance spectroscopy in vivo.

We describe a patient with multiple cerebral lesions that initially were believed to be tumor metastases but were later found to be caused by Nocardia asteroides, after magnetic resonance spectroscopy (MRS) findings suggested that the lesions were infective in origin. This case report demonstrates the ability of MRS, a noninvasive imaging modality, to aid in the diagnosis of brain infection.

Aged↗

PubMed2002

Isolation and characterisation of the phospholipase B gene of Cryptococcus neoformans var. gattii.

Cryptococcus neoformans var. gattii (serotypes B and C) is a human pathogen, ecologically, biochemically, clinically and genetically different from C. neoformans var. grubii (serotype A) and C. neoformans var. neoformans (serotype D). The phospholipase B (PLB1) gene from serotypes B and C was isolated and characterised. It resembled the serotype A and D genes, with an overall sequence homology of more than 85%. The respective open reading frames were 2236 bp (serotype B) and 2239 bp (serotype C) in length. Each contained six introns and encoded a 68-kDa protein destined for secretion. PLB1 was located on the second smallest chromosome in both serotypes. Gene expression, measured as mRNA, was not regulated by temperature, pH or exogenous nutrients.

Amino Acid Sequence↗

PubMed2000

A rapid method for detecting extracellular proteinase activity in Cryptococcus neoformans and a survey of 63 isolates.

A rapid method to detect extracellular proteolytic activity around colonies of Cryptococcus neoformans was developed with tannic acid used to complex with residual protein in a solid medium. A survey was conducted with 32 isolates of C. neoformans var. gattii and 31 isolates of C. neoformans var. neoformans which were cultured on medium containing gelatin as the sole nitrogen source. The annulus of clearing around fungal colonies was > 1.2 mm in 24 (77%) isolates of C. neoformans var. neoformans compared with only 7 (22%) isolates of C. neoformans var. gattii. There was no difference in proteolytic activity between environmental and human clinical isolates of C. neoformans. However, there was a difference between the size of the annulus around animal isolates of C. neoformans var. neoformans and isolates of the same variety from other sources. The annuli around the 14 animal isolates were all >1.2 mm, while 7 (70%) of 10 human clinical isolates and only 3 (43%) of 7 environmental isolates were scored in the high proteinase range. A difference between the genetic types (as characterised by RAPD typing) of C. neoformans var. gattii was also evident with 17 (77%) of 22 VG-I isolates having a small annulus compared with only 1 (17%) of 6 VG-II and VG-III isolates with annuli of similar size. Relatively low proteinase production by C. neoformans var. gattii may reduce local and systemic spread of infection in mammalian hosts.

Animals↗

PubMed1999

Biochemical and functional characterisation of secreted phospholipase activities from Cryptococcus neoformans in their naturally occurring state.

A recent study demonstrated that phospholipase B (PLB), lysophospholipase (LPL) and lysophopholipase transacylase (LPTA) are secreted by Cryptococcus neoformans var. neoformans and showed that the amount of enzyme production correlated with virulence in mice. The present study characterised the extracellular enzyme activities further by radiometric assays and 31P nuclear magnetic resonance spectroscopy (NMR). All three enzymes were most active between 25 and 40 degrees C. Bovine lung surfactant and its major lipid components, disaturated phosphatidylcholine and phosphatidylglycerol, were the optimal substrates for PLB. Lysophosphatidylcholine was the favoured substrate for LPL and LPTA. PLB and LPL/LPTA were differentially affected by Triton X-100, and palmitoyl carnitine was a potent inhibitor of the three phospholipases. LPL and PLB activities were inhibited by dithiothreitol; N-ethylmaleimide inhibited LPL and LPTA activities. None of the enzymes was inhibited by N-bromosuccinimide or p-bromophenacyl bromide. Cellular disruption experiments indicated that >85% of the phospholipase activities were cell-associated, with LPL and LPTA being more easily released than PLB. At pH 5.5 and 7.0, the heat-inactivated secreted enzyme preparations decreased the viability of human neutrophils. This effect was attenuated by active supernates. The relative activities of the PLB, LPL and LPTA in the environment of neutrophils are likely to determine the fate of these cells in vivo. Both phospholipases and heat-stable substances secreted by C. neoformans at 37 degrees C could contribute to membrane degradation and virulence.

Cell Survival↗