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J Kreft

Publications and source records attributed to J Kreft.

At least 19 recordsLinked to original sources

Onset of patterns in an oscillated granular layer: continuum and molecular dynamics simulations.

We study the onset of patterns in vertically oscillated layers of frictionless dissipative particles. Using both numerical solutions of continuum equations to Navier-Stokes order and molecular dynamics (MD) simulations, we find that standing waves form stripe patterns above a critical acceleration of the cell. Changing the frequency of oscillation of the cell changes the wavelength of the resulting pattern; MD and continuum simulations both yield wavelengths in accord with previous experimental results. The value of the critical acceleration for ordered standing waves is approximately 10% higher in molecular dynamics simulations than in the continuum simulations, and the amplitude of the waves differs significantly between the models. The delay in the onset of order in molecular dynamics simulations and the amplitude of noise below this onset are consistent with the presence of fluctuations which are absent in the continuum theory. The strength of the noise obtained by fit to Swift-Hohenberg theory is orders of magnitude larger than the thermal noise in fluid convection experiments, and is comparable to the noise found in experiments with oscillated granular layers and in recent fluid experiments on fluids near the critical point. Good agreement is found between the mean field value of onset from the Swift-Hohenberg fit and the onset in continuum simulations. Patterns are compared in cells oscillated at two different frequencies in MD; the layer with larger wavelength patterns has less noise than the layer with smaller wavelength patterns.

Journal Article↗

Crucial role of sidewalls in velocity distributions in quasi-two-dimensional granular gases.

Our experiments and three-dimensional molecular dynamics simulations of particles confined to a vertical monolayer by closely spaced frictional walls (sidewalls) yield velocity distributions with non-Gaussian tails and a peak near zero velocity. Simulations with frictionless sidewalls are not peaked. Thus interactions between particles and their containers are an important determinant of the shape of the distribution and should be considered when evaluating experiments on a constrained monolayer of particles.

Journal Article↗

Mathematical modelling of biofilm structures.

The morphology of biofilms received much attention in the last years. Several concepts to explain the development of biofilm structures have been proposed. We believe that biofilm structure formation depends on physical as well as general and specific biological factors. The physical factors (e.g. governing substrate transport) as well as general biological factors such as growth yield and substrate conversion rates are the basic factors governing structure formation. Specific strain dependent factors will modify these, giving a further variation between different biofilm systems. Biofilm formation seems to be primarily dependent on the interaction between mass transport and conversion processes. When a biofilm is strongly diffusion limited it will tend to become a heterogeneous and porous structure. When the conversion is the rate-limiting step, the biofilm will tend to become homogenous and compact. On top of these two processes, detachment processes play a significant role. In systems with a high detachment (or shear) force, detachment will be in the form of erosion, giving smoother biofilms. Systems with a low detachment force tend to give a more porous biofilm and detachment occurs mainly by sloughing. Biofilm structure results from the interplay between these interactions (mass transfer, conversion rates, detachment forces) making it difficult to study systems taking only one of these factors into account.

Bacteria↗

Comparative genomics of Listeria species.

Listeria monocytogenes is a food-borne pathogen with a high mortality rate that has also emerged as a paradigm for intracellular parasitism. We present and compare the genome sequences of L. monocytogenes (2,944,528 base pairs) and a nonpathogenic species, L. innocua (3,011,209 base pairs). We found a large number of predicted genes encoding surface and secreted proteins, transporters, and transcriptional regulators, consistent with the ability of both species to adapt to diverse environments. The presence of 270 L. monocytogenes and 149 L. innocua strain-specific genes (clustered in 100 and 63 islets, respectively) suggests that virulence in Listeria results from multiple gene acquisition and deletion events.

Adaptation, Physiological↗

Pathogenicity islands and virulence evolution in Listeria.

As in other bacterial pathogens, the virulence determinants of Listeria species are clustered in genomic islands scattered along the chromosome. This review summarizes current knowledge about the structure, distribution and role in pathogenesis of Listeria virulence loci. Hypotheses about the mode of acquisition and evolution of these loci in this group of Gram-positive bacteria are presented and discussed.

Amino Acid Sequence↗

Regulation of virulence genes in Listeria.

As in all pathogenic bacteria, virulence of the facultative intracellular Listeria species is a multifactorial trait. The expression of the bacterial genes involved in the different steps of the infectious process--invasion, intracellular multiplication and spreading--is temporally and spatially controlled, thus ensuring the presence of the respective gene products at the right moment and place. So far, one network which is involved in the regulation of listerial virulence, the PrfA regulon, has been characterized rather well. The key element of this regulon, PrfA, belongs to the Crp/Fnr family of transcriptional regulators. Its synthesis and activity are influenced by a variety of physico-chemical signals outside and inside of eukaryotic host cells. The analysis of virulence gene expression in vivo, i.e. in infected host cells, indicates that yet uncharacterized bacterial factors other than PrfA, and possibly also host factors, modulate the expression of the PrfA regulon. Essentially nothing is known about the signal transduction pathways involved in the observed differential expression of virulence genes. Fermentable carbon sources seem to have a particular role in virulence gene regulation. In addition to the PrfA regulon, the Clp stress proteins have an impact on Listeria virulence. These two regulons interact with each other by an unknown mechanism.

Bacterial Proteins↗

Listeria pathogenesis and molecular virulence determinants.

The gram-positive bacterium Listeria monocytogenes is the causative agent of listeriosis, a highly fatal opportunistic foodborne infection. Pregnant women, neonates, the elderly, and debilitated or immunocompromised patients in general are predominantly affected, although the disease can also develop in normal individuals. Clinical manifestations of invasive listeriosis are usually severe and include abortion, sepsis, and meningoencephalitis. Listeriosis can also manifest as a febrile gastroenteritis syndrome. In addition to humans, L. monocytogenes affects many vertebrate species, including birds. Listeria ivanovii, a second pathogenic species of the genus, is specific for ruminants. Our current view of the pathophysiology of listeriosis derives largely from studies with the mouse infection model. Pathogenic listeriae enter the host primarily through the intestine. The liver is thought to be their first target organ after intestinal translocation. In the liver, listeriae actively multiply until the infection is controlled by a cell-mediated immune response. This initial, subclinical step of listeriosis is thought to be common due to the frequent presence of pathogenic L. monocytogenes in food. In normal individuals, the continual exposure to listerial antigens probably contributes to the maintenance of anti-Listeria memory T cells. However, in debilitated and immunocompromised patients, the unrestricted proliferation of listeriae in the liver may result in prolonged low-level bacteremia, leading to invasion of the preferred secondary target organs (the brain and the gravid uterus) and to overt clinical disease. L. monocytogenes and L. ivanovii are facultative intracellular parasites able to survive in macrophages and to invade a variety of normally nonphagocytic cells, such as epithelial cells, hepatocytes, and endothelial cells. In all these cell types, pathogenic listeriae go through an intracellular life cycle involving early escape from the phagocytic vacuole, rapid intracytoplasmic multiplication, bacterially induced actin-based motility, and direct spread to neighboring cells, in which they reinitiate the cycle. In this way, listeriae disseminate in host tissues sheltered from the humoral arm of the immune system. Over the last 15 years, a number of virulence factors involved in key steps of this intracellular life cycle have been identified. This review describes in detail the molecular determinants of Listeria virulence and their mechanism of action and summarizes the current knowledge on the pathophysiology of listeriosis and the cell biology and host cell responses to Listeria infection. This article provides an updated perspective of the development of our understanding of Listeria pathogenesis from the first molecular genetic analyses of virulence mechanisms reported in 1985 until the start of the genomic era of Listeria research.

Animals↗

The bvr locus of Listeria monocytogenes mediates virulence gene repression by beta-glucosides.

The beta-glucoside cellobiose has been reported to specifically repress the PrfA-dependent virulence genes hly and plcA in Listeria monocytogenes NCTC 7973. This led to the hypothesis that beta-glucosides, sugars of plant origin, may act as signal molecules, preventing the expression of virulence genes if L. monocytogenes is living in its natural habitat (soil). In three other laboratory strains (EGD, L028, and 10403S), however, the effect of cellobiose was not unique, and all fermentable carbohydrates repressed hly. This suggested that the downregulation of virulence genes by beta-glucosides is not a specific phenomenon but, rather, an aspect of a global regulatory mechanism of catabolite repression (CR). We assessed the effect of carbohydrates on virulence gene expression in a panel of wild-type isolates of L. monocytogenes by using the PrfA-dependent phospholipase C gene plcB as a reporter. Utilization of any fermentable sugar caused plcB repression in wild-type L. monocytogenes. However, an EGD variant was identified in which, as in NCTC 7973, plcB was only repressed by beta-glucosides. Thus, the regulation of L. monocytogenes virulence genes by sugars appears to be mediated by two separate mechanisms, one presumably involving a CR pathway and another specifically responding to beta-glucosides. We have identified in L. monocytogenes a 4-kb operon, bvrABC, encoding an antiterminator of the BglG family (bvrA), a beta-glucoside-specific enzyme II permease component of the phosphoenolpyruvate-sugar phosphotransferase system (bvrB), and a putative ADP-ribosylglycohydrolase (bvrC). Low-stringency Southern blots showed that this locus is absent from other Listeria spp. Transcription of bvrB was induced by cellobiose and salicin but not by arbutin. Disruption of the bvr operon by replacing part of bvrAB with an interposon abolished the repression by cellobiose and salicin but not that by arbutin. Our data indicate that the bvr locus encodes a beta-glucoside-specific sensor that mediates virulence gene repression upon detection of cellobiose and salicin. Bvr is the first sensory system found in L. monocytogenes that is involved in environmental regulation of virulence genes.

Bacterial Proteins↗

A novel PrfA-regulated chromosomal locus, which is specific for Listeria ivanovii, encodes two small, secreted internalins and contributes to virulence in mice.

Several large, cell wall-associated internalins and one small, secreted internalin (InlC) have been described previously in Listeria monocytogenes. Using degenerate primers derived from sequenced peptides of an L. ivanovii major secreted protein, we identified a new 4.25 kb internalin locus of L. ivanovii, termed i-inlFE. The two proteins encoded by this locus, i-InlE and i-InlF, belong to the group of small, secreted internalins. Southern blot analyses show that the i-inlFE locus does not occur in L. monocytogenes. These data also indicate that six genes encoding small, secreted internalins are present in L. ivanovii, in contrast to L. monocytogenes, in which inlC encodes the only small internalin. The mature i-InlE protein (198 amino acids) is secreted in large amounts into the brain-heart infusion (BHI) culture medium in the stationary growth phase. In minimum essential medium (MEM), which has been used previously to induce PrfA-dependent gene transcription, i-inlE mRNA and i-InlE protein are expressed at high levels. As shown by Northern blot analysis and primer extension, transcription of the tandemly arranged i-inlF and i-inlE genes is dependent on the virulence regulator PrfA, and characteristic palindromic sequences ('PrfA-boxes') were identified in the promoter regions of i-inlF and i-inlE. Non-polar i-inlE and i-inlF deletion mutants and an i-inlFE double deletion mutant were constructed and tested in the mouse infection model. After intravenous infection, all three mutants entirely failed to kill C57BL/6 mice even at high infectious doses of 109 bacteria per mouse, whereas the LD50 for the parental strain was determined as 4 x 107 bacteria per mouse. These data suggest an important role for i-InlE and i-InlF in L. ivanovii virulence.

Amino Acid Sequence↗

Complementation of Listeria seeligeri with the plcA-prfA genes from L. monocytogenes activates transcription of seeligerolysin and leads to bacterial escape from the phagosome of infected mammalian cells.

Infection experiments have shown that the avirulent species Listeria seeligeri invaded the enterocyte-like cell line Caco-2 with low efficiency but was unable to escape from the phagosome. Introduction of the listeriolysin gene (hly) from L. monocytogenes into L. seeligeri via a recombinant plasmid did not change these characteristics. No measurable transcription of this gene or of the structurally intact chromosomal seeligerolysin gene (lso) was detected. Transformation with a plasmid carrying the bicistronically transcribed plcA-prfA genes from L. monocytogenes resulted in the efficient expression of the plasmid-encoded transcription activator PrfA, a readily detectable synthesis of seeligerolysin and the escape of the bacteria from the phagosome of infected mammalian cells, followed by intracytoplasmic multiplication.

Animals↗

Physical activity, change in blood pressure and predictors of mortality in older South Africans--a 2-year follow-up study.

OBJECTIVE: A 2-year follow-up study of a cohort of 200 historically disadvantaged older South Africans was conducted to: (i) characterise current levels of habitual physical activity; (ii) relate physical activity to current risk factors for chronic disease; and (iii) identify risk factors associated with 2-year mortality. The baseline sample, drawn in 1993, was found to have a high prevalence of hypertension (71.7%). RESEARCH DESIGN: Retrospective cohort study. METHODS: A baseline sample of 200 persons aged > or = 65 years, resident in the Cape Peninsula, was randomly drawn by means of a two-stage cluster design. Baseline measurements included: anthropometry, waist/hip ratio, systolic and diastolic blood pressure, body mass index (BMI), serum albumin, serum ferritin, haemoglobin and fasting plasma glucose levels, plasma lipid profiles, oral glucose tolerance test and self-reported health status. Subjects were revisited after 2 years, at which time an adapted version of the Yale Physical Activity Survey was administered and measurements of blood pressure and anthropometry were repeated. STATISTICAL ANALYSES: Spearman's rank-order correlations were used to describe relationships between various current risk factors and physical activity. Logistic regression was used to determine predictors of 2-year mortality from baseline data. RESULTS: At follow-up, 142 of the subjects (66 men, 76 women) were traced and measurements collected. Thirty-two subjects were reported to have died by relatives living in the same household (22 men, 10 women). Levels of reported physical activity in the survivors were two-thirds lower than those reported in a sample of North Americans of similar age. There was an inverse association between age and physical activity (r = -0.31; P < 0.0005) and a positive association between BMI and physical activity (r = 0.29; P < 0.005). There was, however, no association between physical activity and systolic or diastolic blood pressure. In men, BMI in the lower tertile (P = 0.07) and serum ferritin levels were positively associated with increased mortality. Serum albumin levels were protective over the 2-year follow-up period (OR = 0.85; P < 0.05). In women, being diabetic (OR = 4.88; P = 0.06) and having a waist/hip ratio in the upper tertile (OR = 3.26; P = 0.06) were associated with mortality. CONCLUSIONS: Physical activity levels in this sample of older historically disadvantaged South Africans were habitually low. Simple anthropometric assessments incorporating weight and waist/hip ratio, together with serum albumin measurements, may be useful to screen general health risk for older adults at primary care level and provide indications for social or medical intervention. Further, strategies for earlier detection and effective management of diabetes, particularly in older women, may reduce premature mortality in this population.

Aged↗

Regulation of virulence gene expression in pathogenic Listeria.

Dynamic interactions between host and pathogen are characteristic of infections caused by intracellular bacteria. This has favoured the evolution of highly effective control systems by which these pathogens regulate the expression of different virulence factors during sequential steps of the infection process. In the case of the facultative intracellular bacterium Listeria monocytogenes, these steps involve internalization by eukaryotic cells, lysis of the resulting phagosome, replication as well as movement within the host cytoplasm, direct cell-to-cell spread, and subsequent lysis of a double-membrane vacuole when entering neighbouring cells. Virulence factors which are involved in each of these steps have been identified and the expression of these factors is subject to a co-ordinate and differential control exerted by the major listerial virulence regulator PrfA. This protein belongs to the Crp/Fnr-family of transcriptional activators and recognizes specific target sequences in promoter regions of several listerial virulence genes. Differential expression of these genes during sequential steps of the infection seems to be at least partially mediated by different binding affinities of PrfA to its target sequences. Activity of PrfA-dependent genes and of prfA itself is under the control of several environmental variables which are used by the pathogen to recognize its transition from the free environment into a eukaryotic host.

Animals↗

The actin-polymerization protein from Listeria ivanovii is a large repeat protein which shows only limited amino acid sequence homology to ActA from Listeria monocytogenes.

Within infected eukaryotic cells the two pathogenic Listeria species, L. monocytogenes and L. ivanovii, induce polymerization of cellular actin and the formation of a propulsive actin tail at one bacterial pole. For L. monocytogenes it has been shown that the product of the listerial actA gene is required for this process which is regarded as a model for actin-based motility. We have now cloned and sequenced a functionally analogous gene from L. ivanovii; its product, as deduced from the DNA sequence, is considerably larger (108 kDa) than L. monocytogenes ActA (67 kDa) and shares only a limited amino acid sequence homology (46% similarity on average) with the latter protein. This is the first example of a virulence gene product from L. ivanovii which is significantly different from its L. monocytogenes counterpart. Comparison of the two ActA proteins gives new insight into the structure of this class of actin-polymerization proteins, in particular with respect to their proline-rich repeat region.

Amino Acid Sequence↗

The virulence regulator protein of Listeria ivanovii is highly homologous to PrfA from Listeria monocytogenes and both belong to the Crp-Fnr family of transcription regulators.

The two pathogenic Listeria species, L. ivanovii and L. monocytogenes, can be differentiated biochemically and show different host ranges. Virulence of L. monocytogenes is dependent on the integrity of prfA which positively and co-ordinately regulates transcription of several virulence genes. Until now, a prfA homologue had not been identified in L. ivanovii. We have now cloned a chromosomal region from L. ivanovii comprising two genes with high homology to the plcA and prfA genes from L. monocytogenes. Distal from prfA, an open reading frame highly homologous to a phosphoribosyl pyrophosphate synthetase gene (prs) was newly identified, defining the border of the virulence gene cluster. Transcription of the gene for ivanolysin O and expression of other genes of the virulence gene cluster in L. ivanovii were dependent on PrfA. The pattern of PrfA-dependent proteins (PdPs) expressed in L. ivanovii was similar, but not identical to that of L. monocytogenes. The PrfA proteins, as predicted from nucleotide sequences of both pathogenic Listeria species, are very similar and show significant homology to the Crp-Fnr family of global transcription regulators.

Amino Acid Sequence↗

A gene encoding a superoxide dismutase of the facultative intracellular bacterium Listeria monocytogenes.

A gene (lmsod) encoding superoxide dismutase (SOD; EC 1.15.1.1) of the facultative intracellular pathogen, Listeria monocytogenes, was cloned by functional complementation of an SOD-deficient Escherichia coli mutant. The nucleotide sequence was determined and the deduced amino acid (aa) sequence (202 aa) showed close similarity to manganese-containing SOD's from other organisms. Subunits of the recombinant L. monocytogenes SOD (re-SOD) and of both E. coli SODs formed enzymatically active hybrid enzymes in vivo. DNA/DNA-hybridization experiments showed that this type of recombinant re-sod gene is conserved within the genus Listeria.

Amino Acid Sequence↗